Electrophysiological classes of layer 2/3 pyramidal cells in

monkey prefrontal cortex

A. V. Zaitsev, N. V. Povysheva, G. Gonzalez-Burgos and D. A. Lewis
J Neurophysiol 108:595-609, 2012. First published 11 April 2012;
doi: 10.1152/jn.00859.2011

You might find this additional info useful...

This article cites 89 articles, 56 of which you can access for free at:
http://jn.physiology.org/content/108/2/595.full#ref-list-1
This article has been cited by 1 other HighWire-hosted articles:
http://jn.physiology.org/content/108/2/595#cited-by

Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
Updated information and services including high resolution figures, can be found at:
http://jn.physiology.org/content/108/2/595.full
Additional material and information about Journal of Neurophysiology can be found at:
http://www.the-aps.org/publications/jn

This information is current as of April 10, 2013.

Journal of Neurophysiology publishes original articles on the function of the nervous system. It is published 24
times a year (twice monthly) by the American Physiological Society, 9650 Rockville Pike, Bethesda MD
20814-3991. Copyright © 2012 the American Physiological Society. ESSN: 1522-1598. Visit our website at
http://www.the-aps.org/.

Electrophysiological classes of layer 2/3 pyramidal cells in monkey
prefrontal cortex
A. V. Zaitsev,1,2 N. V. Povysheva,2,3 G. Gonzalez-Burgos,2 and D. A. Lewis2,3
1
Sechenov Institute of Evolutionary Physiology and Biochemistry of the Russian Academy of Sciences (IEPhB),
Saint-Petersburg, Russia; 2Department of Psychiatry, University of Pittsburgh, Pittsburgh, Pennsylvania;
and 3Department of Neuroscience, University of Pittsburgh, Pittsburgh, Pennsylvania
Submitted 21 September 2011; accepted in final form 10 April 2012
Zaitsev AV, Povysheva NV, Gonzalez-Burgos G, Lewis DA.
Electrophysiological classes of layer 2/3 pyramidal cells in monkey
prefrontal cortex. J Neurophysiol 108: 595– 609, 2012. First published
April 11, 2012; doi:10.1152/jn.00859.2011.—The activity of supra-
granular pyramidal neurons in the dorsolateral prefrontal cortex
(DLPFC) neurons is hypothesized to be a key contributor to the
cellular basis of working memory in primates. Therefore, the intrinsic
membrane properties, a crucial determinant of a neuron’s functional
properties, are important for the role of DLPFC pyramidal neurons in
working memory. The present study aimed to investigate the biophys-
ical properties of pyramidal cells in layer 2/3 of monkey DLPFC to
create an unbiased electrophysiological classification of these cells.
Whole cell voltage recordings in the slice preparation were performed
in 77 pyramidal cells, and 24 electrophysiological measures of their
passive and active intrinsic membrane properties were analyzed.
Based on the results of cluster analysis of 16 independent electro-
physiological variables, 4 distinct electrophysiological classes of
monkey pyramidal cells were determined. Two classes contain regu-
lar-spiking neurons with low and high excitability and constitute 52%
of the pyramidal cells sampled. These subclasses of regular-spiking
neurons mostly differ in their input resistance, minimum current that
evoked firing, and current-to-frequency transduction properties. A
third class of pyramidal cells includes low-threshold spiking cells
(17%), which fire a burst of three-five spikes followed by regular
firing at all suprathreshold current intensities. The last class consists of
cells with an intermediate firing pattern (31%). These cells have two
modes of firing response, regular spiking and bursting discharge,
depending on the strength of stimulation and resting membrane
potential. Our results show that diversity in the functional properties
of DLPFC pyramidal cells may contribute to heterogeneous modes of
information processing during working memory and other cognitive
operations that engage the activity of cortical circuits in the superficial
layers of the DLPFC.
firing pattern; intrinsic membrane properties; low-threshold spiking;
regular spiking
THE PREFRONTAL CORTEX (PFC), one of the largest and most
differentiated neocortical areas of the primate brain, mediates
complex behaviors that are characteristic of primates in general
and of humans in particular (Wood and Grafman 2003). Work-
ing memory, the ability to hold information in mind for a short
period of time, is a critical cognitive process underlying com-
plex behaviors in primates (Baddeley 2003; Fuster 1997; Gold-
man-Rakic 1995). In vivo, cells in the dorsolateral PFC
(DLPFC) frequently remain active during the delay period of a
delayed-response trial, and thus DLPFC neuron activity is
Address for reprint requests and other correspondence: A. V. Zaitsev,
IEPhB, Toreza prospect, 44, Saint-Petersburg, 194223, Russia (e-mail:
aleksey_zaitsev@mail.ru).
www.jn.org 0022-3077/12 Copyright © 2012 the American Physiological Society
J Neurophysiol 108: 595– 609, 2012.
First published April 11, 2012; doi:10.1152/jn.00859.2011.
Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
considered the cellular correlate of mnemonic events in the
working memory process (Goldman-Rakic 1995; Miller et al.
1996). Although the delay period activity has been reported in
many other cortical and subcortical areas (Curtis and Lee 2010;
Hikosaka et al. 2000; Watanabe and Funahashi 2004), DLPFC
cell firing during the delay period is more resistant to distrac-
tors than in other regions (Artchakov et al. 2009; Miller et al.
1996).
The exact mechanisms underlying sustained delay-related
activity of DLPFC neurons are still largely unclear. The spe-
cific organization of neuronal circuitry in this brain area,
especially in layer 2/3 (Gonzalez-Burgos et al. 2000; Kritzer
and Goldman-Rakic 1995; Melchitzky et al. 1998, 2001; Pucak
et al. 1996; Selemon and Goldman-Rakic 1988), and distinc-
tive morphological and electrophysiological properties of these
neurons may contribute to the unique features of delay-related
firing in PFC. For example, anatomical observations of pyra-
midal cells in human and macaque monkey PFC revealed a
higher number of dendritic branches and higher spine density
than their counterparts in other cortical regions (Elston 2003;
Elston et al. 2001; Jacobs et al. 2001). Such increased dendritic
complexity and spine density of PFC neurons may reflect the
integration of a large number of incoming inputs of diverse
origin. Furthermore, it may also indicate a specialization of
DLPFC neurons in recurrent excitation, a proposed mechanism
for generation of persistent delay-related activity (Wang 2001).
The intrinsic firing properties of pyramidal neurons may be
a crucial determinant of in vivo firing and behavioral perfor-
mance. For instance, age-related changes of intrinsic mem-
brane properties of monkey DLPFC neurons correlate with
age-related changes in working memory performance (Chang
et al. 2005; Luebke et al. 2010; Luebke and Amatrudo 2010)
and thus could explain the age-related changes in working
memory task-related firing of monkey DLPFC neurons in vivo
(Wang et al. 2011). Despite the importance of intrinsic mem-
brane properties of primate DLPFC pyramidal neurons, only a
few studies have reported detailed data on the electrophysio-
logical properties of primate DLPFC pyramidal cells, focusing
mostly on layer 5 pyramidal neurons (Chang and Luebke
2007). We have performed multiple studies of neurons, syn-
apses, and small circuits in layer 3 of monkey DLPFC, ad-
dressing mostly properties of GABA neurons (Gonzalez-
Burgos et al. 2004, 2005a, 2009; Krimer et al. 2005; Povysheva
et al. 2007, 2008; Zaitsev et al. 2005, 2009). Moreover, our
studies of layer 3 pyramidal neurons from monkey DLPFC
were focused on synaptic properties (Gonzalez-Burgos et al.
2004, 2008; Povysheva et al. 2006) and neuromodulation
595
596 MONKEY PFC PYRAMIDAL CELLS
(Gonzalez-Burgos et al. 2005b; Henze et al. 2000; Urban et al.
2002), but no detailed description of the electrophysiological
subclasses of layer 3 pyramidal cells was obtained.
Current classifications of pyramidal cells are based on the
cells’ firing patterns as well as the shape of their action
potentials (Connors and Gutnick 1990; de la Pena and Geijo-
Barrientos 1996; Degenetais et al. 2002; Foehring et al. 1991;
Larkman and Mason 1990; Maravall et al. 2004; Yang et al.
1996), whereas the subthreshold membrane properties are
often overlooked (but see Kawaguchi 1993; Schwindt et al.
1997). Recently, we have shown that multivariate statistical
analysis of a large neuronal data set is a powerful classification
tool that allowed us to reliably define multiple subclasses of
GABA neurons in layer 3 of monkey PFC (Krimer et al. 2005;
Zaitsev et al. 2005, 2009). In the present study we employed
cluster analysis for unbiased delineation of the physiological
groups of pyramidal cells in layer 2/3 of monkey DLPFC.
Sixteen electrophysiological measures of subthreshold, single
spike, and firing pattern properties have been chosen for this
multivariate statistical analysis. We describe four distinct elec-
trophysiological classes of pyramidal cells in DLPFC, includ-
ing two classes of regular spiking (RS) cells with different
input resistance, low-threshold spiking (LTS) cells, and a class
of cells with intermediate properties.
MATERIALS AND METHODS
Slice Preparation
Twelve experimentally naive young adult (3.5– 6 kg; 3.5– 4-years
old), male, long-tailed macaque monkeys (Macaca fascicularis) were
used in this study. All studies were conducted in accordance with the
guidelines of the National Institutes of Health Guide for the Care and
Use of Laboratory Animals and were approved by the University of
Pittsburgh Institutional Animal Care and Use Committee. The proce-
dure used to obtain tissue from the DLPFC has been previously
described in detail (Gonzalez-Burgos et al. 2004). Briefly, animals
were treated with ketamine hydrochloride (25 mg/kg im), dexameth-
asone phosphate (0.5 mg/kg im), and atropine sulfate (0.05 mg/kg sc);
an endotracheal tube was inserted, and anesthesia was maintained
with 1% halothane in a 28% O2-air mixture. Monkeys were placed in
a stereotaxic apparatus, and a craniotomy was performed over the
DLPFC in one hemisphere. The dura was removed in a location
determined by stereotaxic coordinates and by the position of relevant
sulcal landmarks, and a small block of tissue was excised containing
both the medial and lateral banks of the principal sulcus (area 46) as
well as a small adjacent portion of dorsal area 9. After the surgery, the
animals were treated with an antibiotic (chloramphenicol, 15 mg/kg
im) and an analgesic (hydromorphone, 0.02 mg/kg im) three times a
day for 3 days. All animals recovered quickly with no impairments in
eating or drinking and no overt behavioral deficits. In most cases, the
animals underwent the same procedure 2– 4 wk later to obtain tissue
from a nonhomotopic portion of the contralateral areas 46 and 9.
During the second procedure, after the craniotomy, the animal was
given an overdose of pentobarbital sodium (30 mg/kg) and perfused
through the heart with ice-cold modified artificial cerebrospinal fluid
before the tissue block was quickly excised. Subsequent treatment of
the tissue was the same for both procedures. Previous studies have
demonstrated that the initial biopsy did not affect the properties of
neurons obtained in the contralateral hemisphere (Gonzalez-Burgos et
al. 2000, 2004; Henze et al. 2000).
The tissue blocks were placed in ice-cold Ringer solution contain-
ing (in mM) 126 NaCl, 2.5 KCl, 1.25 NaH2PO4, 2 CaCl2, 1 MgSO4,
26 NaHCO3, and 10 dextrose, pH 7.4, perfused with a 95%O2-5%CO2
gas mixture. Coronal 350-␮m-thick slices were cut from each block
J Neurophysiol • doi:10.1152/jn.00859.2011 • www.jn.org
using a Vibratome (VT 1000S Leica, Wetzlar, Germany) and incu-
bated for 1 h at 36°C and at room temperature thereafter, or at room
temperature from the beginning. For recordings, slices were sub-
merged in a chamber mounted on the microscope and perfused with
Ringer solution at 32°C. Other brain slices from these animals were
used and reported in studies of interneurons (Povysheva et al. 2006,
2008; Zaitsev et al. 2005, 2009).
Electrophysiological Recordings
Pyramidal cells in layer 2/3 were visualized using infrared differ-
ential interference contrast (DIC) videomicroscopy and identified on
the basis of their triangular soma and the presence of an apical
dendrite. Patch electrodes with open-tip resistances of 5–10 M⍀ were
Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
filled with a solution containing (in mM) 114 potassium gluconate, 6
KCl, 4 MgATP, 0.3 GTP, 10 HEPES, 0.5% biocytin, and pH 7.25
adjusted with KOH. Whole cell current-clamp recordings were per-
formed after seal resistance of at least 1.5–2 G⍀ was reached.
Voltages were amplified using Intracellular Electrometers IE-210
(Warner Instrument, Hamden, CT) or MultiClamp 700A (Axon In-
struments, Union City, CA) operating in a bridge-balance mode,
filtered online at 4 –5 kHz, and acquired on a personal computer at a
sampling rate of 20 kHz using Power 1401 interface and Signal 2 or
Signal 3 software (CED, Cambridge, UK). Access resistance typically
was 15–30 M⍀ and remained relatively stable during experiments
(ⱕ30% increase) for the cells included in the analysis. Membrane
potential was not corrected for the liquid junction potential. To
characterize the intrinsic membrane properties of neurons, hyper- and
depolarizing current steps of 500-ms duration were applied in 5- to
10-pA increments at 0.2 Hz with two repetitions. Recordings of
intrinsic membrane properties were done immediately after membrane
rupture and lasted less than 5 min. All included cells had resting
potentials more negative than ⫺50 mV and overshooting spikes.
Electrophysiological Analysis
A number of electrophysiological parameters were measured. Rest-
ing membrane potential (RMP; in mV) was determined as the stable
membrane potential (no holding current applied) reached a few
minutes after breaking the membrane.
Subthreshold membrane properties. Input resistance (Rin; in M⍀),
is the slope of the regression line fitted to the voltage-current (V-I)
curve (usually between ⫺50 and ⫺10 pA) as measured at the end of
the 500-ms voltage responses. The membrane time constant ␶ (in ms)
was determined from the monoexponential curve best fitting the
average voltage response to the small hyperpolarizing current steps.
Rheobase (I0; in pA) is the minimum current that evoked firing. Sag
(in %) was measured during a 500-ms hyperpolarizing current step
(50 pA) as the percent change between the most negative membrane
potential (Vhmax) and steady-state membrane potential at the end of
the step (Vhss): (Vhmax ⫺ Vhss)/Vhss ⫻ 100%. Rebound depolarization
(RD; in %) was measured after a 500-ms hyperpolarizing current step
(50 pA) as a maximal positive voltage deflection above RMP (Vrdmax)
and normalized to the amplitude of steady-state voltage deflection
during that hyperpolarizing current step: Vrdmax/Vhss ⫻ 100%. Hump
(in %) was estimated at the depolarizing current step that preceded
spiking as the percent change between the maximal (Vdmax) and
steady-state membrane potential: (Vdmax ⫺ Vdss)/Vdss ⫻ 100%.
Action potential properties. Action potential (AP) properties were
analyzed at I0. Time to first spike (TFS; in ms) is the time from the
beginning of stimulation to the first spike. Action potential threshold
(APT; in mV) is the membrane potential at the point at which the
interpolated rate of voltage rise (dV/dt) reached ⬎10 mV/ms. Action
potential amplitude (APA; in mV) was measured from the threshold to
the peak. Action potential duration (APD; in ms) is the spike width at
its half-amplitude. Amplitude of the afterhyperpolarization (AHP; in
mV) was measured from the APT to the most negative membrane
 MONKEY PFC PYRAMIDAL CELLS
potential after the spike. Amplitude of the fast component of the AHP
(fAHP; in mV) was measured from the APT to the hyperpolarization
peak or, if present, to the onset of the medium component of the AHP
(mAHP). The latter was determined as the crossing point of a marked
slowing in the voltage drop to ⬍5 mV/ms. The depolarizing afterpo-
tential (ADP; in mV) was measured as the positive voltage deflection
between fast and medium AHPs. Amplitude of the mAHP (in mV)
was measured from the peak ADP to the peak AHP. If ADP was
absent, mAHP was measured from the end of the fAHP. The time
interval tAHP (in ms) was measured as the interval between onset of
AHP and the hyperpolarization peak.
Firing pattern properties. To determine firing pattern properties,
two times the I0 was applied, except for the interspike interval
between the first and second spikes at minimal suprathreshold current
(ISI1–2; in ms). Steady-state frequency (fss; in Hz), the reciprocal of
the average of the four to nine interspike intervals (ISIs), was
measured within the last 250 ms of the response to depolarizing
current pulses, where firing frequency remained relatively stable. Late
frequency adaptation (FFAL; in %), is the percent decrease in the
frequency from onset (reciprocal to the 1st ISI) to steady-state fre-
quency. Initial frequency adaptation (FFAI; in %) is the percent
decrease in frequency, measured as the reciprocal of the first and
second ISIs.
Frequency-current relationship properties.y Frequency-current re-
lationship (f-I) properties were calculated for the first and second
instantaneous firing frequency (1/ISI). fn_k (in Hz/pA) is the slope of
linear trend for the instantaneous firing frequency n (1/ISIn) vs.
current (for n ⫽ 1, 2). fn_threshold (in Hz) is the intercept of the
extrapolated f-I linear fit with the frequency axis (for n ⫽ 1, 2).
Histological Processing and Morphological Analysis
After recordings were made, slices were immersed in 4% parafor-
maldehyde in 0.1 M phosphate buffer for 24 –72 h at 4°C and then
cryoprotected (33% glycerol and 33% ethylene glycol in 0.1 M
phosphate buffer) and stored at ⫺80°C. To visualize biocytin, about
one-half of the slices were incubated with streptavidin-Alexa Fluor
633 conjugate (Invitrogen; dilution 1:500) for 24 – 48 h at 4°C in
phosphate buffer containing 0.4% Triton X-100. Pyramidal cells were
imaged using an Olympus Fluoview 500 confocal laser scanning
microscope equipped with a ⫻20/0.80 N.A. oil-immersion objective.
The remaining slices were serially resectioned at 40 –50 ␮m, and then
the sections were treated with 1% H2O2 for 2–3 h at room tempera-
ture, rinsed, and incubated with the avidin-biotin-peroxidase complex
(1:100; Vector Laboratories, Burlingame, CA) in phosphate buffer for
4 h at room temperature. Sections were rinsed, stained with 3,3=-
diaminobenzidine, mounted on gelatin-coated glass slides, dehy-
drated, and coverslipped. Some of these pyramidal neurons were
reconstructed using the Neurolucida tracing system (MicroBright-
Field, Williston, VT).
Statistical Analysis
All statistical tests were performed using Statistica 6.1 software
(StatSoft, Tulsa, OK). Unless otherwise stated, all data are means and
standard error of measurement. To divide pyramidal cells into groups
based on electrophysiological properties, the cluster analysis was
employed, following Ward’s hierarchical clustering algorithm with
Euclidean distance, which reduced cluster size by consecutively
merging data points based on the least possible increase in the
within-group sum of squared deviation (Johnson and Wichern 1998;
Ward 1963). Before the cluster analysis was performed, all variables
were normalized to their z scores. The statistical significance between
group means was tested using ANOVA followed by Fisher’s least
significant difference (LDS) post hoc tests (multiple comparison
tests).
J Neurophysiol • doi:10.1152/jn.00859.2011 • www.jn.org
597
RESULTS
Classification Based on Subthreshold Responses, AP, and
Firing Pattern Properties
Seventy-seven pyramidal cells from 12 monkeys were in-
cluded in this study (4 –12 neurons per animal). Neurons were
identified as pyramidal cells based on biocytin labeling follow-
ing electrophysiological recording; in each case, these cells had
clear apical and basal dendrites that were densely covered with
spines. The somata of cells were located across the depth of
layer 2/3 (between 300 and 800 ␮m from the pial surface).
Representative examples of reconstructed layer 2/3 pyramidal
cells are shown in Fig. 1.
Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
According to the current qualitative electrophysiological
classifications, pyramidal cells can be grouped into cells that
fire bursts of multiple spikes [intrinsic bursting neurons, also
termed LTS cells] and RS cells that fire trains of single spikes
during current pulse injection; RS cells are a predominant
electrophysiological type of pyramidal neurons in layer 2/3
(Chang and Luebke 2007; de la Pena and Geijo-Barrientos
1996; Degenetais et al. 2002; Foehring et al. 1991; Hattox and
Nelson 2007; Tasker et al. 1996; Yang et al. 1996). Different
subclasses of RS pyramidal cells are described based on
somewhat arbitrary criteria such as the degree of changes in AP
threshold and spike frequency adaptation across a sustained
train of APs (Agmon and Connors 1992; Chang and Luebke
2007; Degenetais et al. 2002; Nunez et al. 1993); however, the
tentative correspondence between groups identified in previous
studies is not always obvious (Degenetais et al. 2002).
In our view, a reliable electrophysiological classification of
pyramidal cells should be based on the most functionally
relevant aspects of a neuron’s electrophysiological properties.
Recently, we showed that a multivariate statistical approach to
the classification of cortical interneurons is superior to “sub-
jective” analyses, because it permits multiple parameters to be
considered in concert, which is very difficult to accomplish
with any type of subjective analysis (Krimer et al. 2005;
Zaitsev et al. 2005). Thus, to determine which electrophysio-
logical classes of pyramidal neurons could be segregated using
a multivariate statistical approach, we performed a cluster
analysis. First, we determined which variables should be in-
cluded. Because a neuron’s functions depend on its 1) sub-
threshold responses, 2) AP, and 3) firing pattern properties, we
chose 24 electrophysiological variables in an effort to compre-
hensively describe these properties (see MATERIALS AND METH-
ODS for details). It is important that selected variables are
independent and describe different parameters of neuronal
behavior. Therefore, we calculated correlation coefficients to
determine whether any of the measures were correlated. Weak-
to-moderate correlations (r ⫽ 0.20 – 0.40) were found between
many variables; however, stronger correlations (r ⬎ 0.50) were
observed only between a few variables. For example, strong
correlations were found between Rin and I0 (r ⫽ ⫺0.73),
between sag, hump, and RD (r ⫽ 0.66 – 0.86), between f1_k,
f2_k, and fss (r ⫽ 0.56 – 0.73), and between f1_threshold and
f2_threshold (r ⫽ 0.72). Of pairs of strongly correlated parame-
ters, only one was included into cluster analysis. Instead of sag,
hump, and RD, we used the derivative parameter equal to the
sum of values of sag, hump, and RD. In total, 16 variables were
chosen, namely, Rin, ␶, APT, APA, APD, AHP, mAHP, ADP,
tAHP, fss, FFAL, FFAI, ISI1–2, f1_k, f1_threshold, (sag ⫹ hump ⫹
598 MONKEY PFC PYRAMIDAL CELLS

Fig. 1. Morphological properties of monkey

dorsolateral prefrontal cortex (DLPFC) pyra-
midal cells. A: lateral view (left) of the ma-
caque monkey cortex showing the approxi-
mate location of the DLPFC tissue blocks
removed during surgery (see MATERIALS AND
METHODS) and schematic view (right) of a
typical coronal slice containing portions of
DLPFC areas 9 and 46, including the medial
and lateral banks of the principal sulcus (PS).

Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
Representative examples are confocal (B) and
Neurolucida-reconstructed layer 2/3 pyrami-
dal cells (C). Labels above cells indicate the
experimental identification number of cells
and, in parentheses, their electrophysiological
type based on subjective classification: RS,
regular spiking; IM, intermediate; LTS, low-
threshold spiking. Cluster numbers are based
on the cluster solutions obtained with Ward’s
clustering method (see text for details). Note
that although the majority of layer 2/3 pyra-
midal cell apical tufts reach the pial surface
and begin to branch exuberantly at the layer
2/1 junction, some pyramidal cells of layer 2/3
resemble slender pyramidal cells from layer 5.
Scale is the same for B and C.

RD); the mean of absolute correlation coefficients between the
16 variables was 0.21 ⫾ 0.01 (P ⬎ 0.05). All variables were
normalized, and then Ward’s hierarchical clustering algorithm
with Euclidean distance was used for classification. The ob-
tained hierarchical tree suggested four main electrophysiolog-
ical classes of pyramidal cells (Fig. 2A).
Next, we checked whether the cells in each cluster had firing
properties that matched some of the properties proposed in
previous studies (Fig. 2B). We found that clusters 1 (n ⫽ 27)
and 2 (n ⫽ 13) included mostly RS cells. These cells generated
trains of single spikes with relatively low frequencies and
prominent frequency adaptation (Fig. 3A). Both the firing
frequency and the frequency adaptation almost invariably in-
creased with an increase in intensity of the stimulating current.
Cluster 4 (n ⫽ 13) consisted mostly of LTS cells, which
discharged with a burst of two to five fast spikes in response to
just suprathreshold depolarizing current steps (Fig. 3B). During
the initial burst, AP threshold slightly increased and the am-
plitude slightly decreased. After the initial spike burst, LTS
neurons fired single spikes with progressive spike frequency
adaptation and stable spike threshold and spike amplitude.
Cluster 3 (n ⫽ 24) included RS cells (n ⫽ 9) and pyramidal
cells (n ⫽ 15), which exhibited firing properties intermediate
(IM) between RS and LTS types (Fig. 3C). The same cell
might exhibit a RS pattern at just above firing threshold
stimulation but show a pattern more similar to that of LTS cells
at an increased level of depolarization current.
The result of the cluster analysis indicates the degree of
similarity among neurons but does not assign statistical signif-
icance to the grouping. To determine whether the clusters
J Neurophysiol • doi:10.1152/jn.00859.2011 • www.jn.org
contained cells with statistically distinct properties, we applied
the ANOVA test to all the measured variables. For any pair of
clusters, the Fisher’s LSD post hoc test was thereafter applied
to compare the means (Table 1). ANOVA revealed significant
differences between clusters for 21 of 24 electrophysiological
variables, confirming the statistical significance of the obtained
classification.
Analyzing the results of the Fisher’s LSD post hoc test, we
found that cells from cluster 4 had the most distinctive prop-
erties because they were different from cells in the other
clusters in the maximal number of electrophysiological prop-
erties (9 of 24), including almost all single-spike properties and
f-I relationship properties. Cluster 3 was different from all
other clusters in its subthreshold properties (Rin, sag), different
kinetics of afterpotentials, and firing frequency adaptation.
Clusters 1 and 2, both consisting of RS cells, differed from
each other in Rin, f-I relationship properties, and some other
properties (Table 1). On the basis of electrophysiological
properties of cells included in each cluster, we named the
clusters accordingly: 1) RS cells with low input resistance
(RS_LRi), 2) RS cells with high input resistance (RS_HRi),
3) IM cells, and 4) LTS cells. Below we provide a compre-
hensive comparison of pyramidal cells from each cluster.
Subthreshold Membrane Properties
RMP was the only parameter that did not differ between
electrophysiological classes of pyramidal cells (ANOVA, F ⫽
0.5, P ⫽ 0.70). The resting potential had a mean value of
72.2 ⫾ 0.6 mV for the total pyramidal cell population follow-
 MONKEY PFC PYRAMIDAL CELLS
Fig. 2. Cluster analysis-based physiological classification of pyramidal neurons
in monkey DLPFC. A: the dendrogram obtained through Ward’s hierarchical
clustering algorithm revealed 4 clusters, demonstrating the presence of distinct
physiological groups of pyramidal neurons. RS_LRi, RS cells with low input
resistance; RS_HRi, RS cells with high input resistance. B: the number of
neurons of each subjectively characterized electrophysiological type in the 4
clusters obtained with Ward’s clustering method. Clusters 1 and 2 included
mostly RS cells, cluster 3 included IM and RS cells, and cluster 4 consisted of
LTS cells.
ing a normal distribution (Shapiro-Wilk’s W-test, W ⫽ 0.98,
P ⫽ 0.22; Kolmogorov-Smirnov test, d ⫽ 0.08, P ⬎ 0.20).
Rin showed remarkable differences between the electrophys-
iological classes. A Fisher’s LSD test revealed that pyramidal
cell classes could be subdivided into three distinct groups
(Table 1). The lowest Rin was exhibited by IM cells (118 ⫾ 7
M⍀), whereas LTS cells showed relatively large Rin (261 ⫾ 42
M⍀). RS_LRi cells had almost two times smaller Rin than
RS_HRi cells (174 ⫾ 16 vs. 332 ⫾ 28 M⍀; Fig. 4). As a
consequence, RS_HRi and LTS cells had lower I0 than
RS_LRi and IM neurons (Table 1). Membrane time constant ␶
was maximal in LTS cells, and thus this cell type may be more
efficient at summation of excitatory synaptic potentials.
In response to hyperpolarizing current steps, IM pyramidal
cells (cluster 3) exhibited a large time-dependent rectification
that produced a delayed depolarizing sag, shifting the mem-
brane potential toward the RMP. The sag was more pro-
nounced when the cells were hyperpolarized more than 10 –20
mV relative to the RMP, suggesting it was likely produced by
the current through hyperpolarization-activated and cyclic nu-
cleotide-gated channels (Ih; Pape 1996; Robinson 2003).
Whereas pyramidal cells from cluster 3 exhibited the largest
sag (20.2 ⫾ 2.6%), cells in the other clusters typically showed
no detectable sag or a small sag (7–13%) (Figs. 4 and 5).
The majority of pyramidal cells that exhibited profound
depolarizing sag during hyperpolarizing current steps showed a
transient RD after the end of the hyperpolarizing current. The
strong correlation between sag and RD (r ⫽ 0.86, P ⬍ 0.001;
J Neurophysiol • doi:10.1152/jn.00859.2011 • www.jn.org
599
Fig. 5A) indicates that the ionic mechanisms of both phenom-
ena are similar and that Ih can contribute to the RD, as well. In
contrast to previous reports (de la Pena and Geijo-Barrientos
1996; Degenetais et al. 2002; Yang et al. 1996), the RD in our
conditions never caused rebound spikes in IM cells, which had
the largest sag amplitude, or in other clusters.
In response to subthreshold depolarizing current steps, al-
most all LTS cells and IM cells exhibited a humplike depolar-
izing voltage deflection, which was not observed in either class
of RS cells (Figs. 4 and 5). The initial bursts of APs in LTS
neurons suggest the involvement of low-threshold calcium
channels underlying the bursting mechanism. In agreement
with this, the amplitude of the humplike depolarization was
Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
voltage dependent in that it increased as the depolarizing
current step was applied from more hyperpolarized membrane
potentials (Fig. 5C). In the total population of cells, the hump
amplitude was significantly correlated with the amplitude of
the sag (r ⫽ 0.66, P ⬍ 0.01; Fig. 5B) and of the rebound
depolarization (r ⫽ 0.67, P ⬍ 0.001; Fig. 5A), indicating that
Ih may contribute to the hump, as well. It is worth noting that
the slope of the regression line for sag vs. hump differed
between pyramidal cell classes (Fig. 5B): LTS cells had the
steepest slope, and although LTS cells had smaller sag than IM
cells (LTS sag: 13.2 ⫾ 2.0%; IM sag: 20.2 ⫾ 2.6%), the hump
amplitude was similar (LTS hump: 32.3 ⫾ 4.6%; IM hump:
31.2 ⫾ 3.7%).
Single-Spike Properties
The properties of individual APs were evaluated from the
first spikes evoked by minimal suprathreshold current. The
latency to the first spike did not differ between cell groups,
with an average value of 131 ⫾ 6 ms. APT was considerably
lower in LTS (⫺44.0 ⫾ 0.8 mV) than in RS pyramidal cells
(⫺40.4 ⫾ 0.9 mV in RS_LRi and ⫺40.0 ⫾ 0.9 mV in
RS_HRi), indicating a greater excitability of the LTS cell class.
LTS cells had the longest spike duration at half-maximal
amplitude (1.12 ⫾ 0.11 ms) compared with other pyramidal
cell classes (0.88 – 0.93 ms). The amplitude of the first spike
was significantly smaller in RS_LRi pyramidal cells (64 ⫾ 4
mV) than in all other cell classes.
In the majority of neurons, the first AP in a train was
followed by the same sequence of afterpotentials; a fast AHP
was followed by a depolarizing afterpotential and then by a
medium AHP. However, the depolarizing afterpotential ampli-
tude varied significantly between cells and was undetectable in
some neurons (Fig. 6). The depolarizing afterpotential was
generally more prominent in IM cells (2.4 ⫾ 0.4 mV) than in
RS cells (0.8 ⫾ 0.2 mV in RS_LRi and 0.5 ⫾ 0.3 mV in
RS_HRi). The medium AHP was larger in amplitude and
almost twice as long in duration in RS_LRi than in RS_HRi
cells. In LTS cells a prominent depolarizing afterpotential led
to the next spike, which completely masked a medium-com-
ponent AHP. After the initial burst of spikes, LTS cells fired
single spikes with the same sequence of afterpotentials as RS
pyramidal cells.
Firing Pattern Properties
Onset response. The most remarkable electrophysiological
feature of LTS cells compared with other cell groups is their
ability to fire bursts of APs at the beginning of the response to
600 MONKEY PFC PYRAMIDAL CELLS
Fig. 3. Representative examples of layer 2/3 pyrami-
dal cell firing pattern from monkey PFC. A–C: subse-
quent sweeps with spikes beginning with the response
to the first suprathreshold current step recorded from
the typical RS (cluster 1) (A), LTS (cluster 4) (B), and
IM (cluster 3)pyramidal cells (C).
just suprathreshold depolarizing current pulses. A typical layer
2/3 LTS pyramidal cell fired just one burst consisting of three
to four spikes followed by a train of single spikes. In contrast,
both classes of RS and IM cells typically generated trains of
single spikes with low frequencies in response to just suprath-
reshold depolarizing current pulses. To quantify this aspect of
firing pattern, the ISI1–2 in the train was selected. ISI1–2 was
measured at just suprathreshold depolarizing current pulses.
We found that LTS cells exhibited the shortest ISI1–2 (27 ⫾ 15
ms), whereas the mean value of ISI1–2 in other pyramidal cell
groups was several times larger (Fig. 7, Table 1).
Firing frequency adaptation. To characterize the firing fre-
quency adaptation, the instantaneous spike frequency was
plotted against the ISI number. With near-threshold depolariz-
ing current steps, both classes of RS cells displayed a non-
adapting or weakly adapting firing pattern, whereas the LTS
cells exhibited strong firing frequency adaptation (Fig. 8). An
increase of current enhanced firing frequency adaptation in RS
and IM cells. In RS pyramidal cells, firing frequency adapta-
tion developed within the first three to six spikes (the first
100 –200 ms after the onset of the current pulses); later, the
firing frequency remained relatively stable until the end of the
current pulse. In response to an increase of current magnitude,
J Neurophysiol • doi:10.1152/jn.00859.2011 • www.jn.org
Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
most IM cells fired a pair of spikes at the beginning of the train.
The initial spike doublet was followed by an almost nonadapt-
ing firing pattern. To compare the degree of adaptation be-
tween cell classes, the firing frequency adaptation was esti-
mated at a current intensity two times the I0. Late and initial
firing frequency adaptations were calculated as the percentage
of decrease in the frequency from the onset instantaneous
frequency (reciprocal of the 1st ISI) to steady-state frequency
(late adaptation) and to the 2nd instantaneous frequency (initial
adaptation). The late adaptation was stronger in LTS (86 ⫾
2%) and IM cells (76 ⫾ 2%) than in RS cells (50 ⫾ 2% in
RS_LRi and 58 ⫾ 5% in RS_HRi). The largest initial firing
frequency adaptation was found for IM cells (66 ⫾ 3%); in all
other pyramidal cell classes, it was only about 40% (Fig. 8E,
Table 1).
Interestingly, the changes in firing frequency adaptation with
increasing current were cell type specific. To estimate this
property, an adaptation ratio was calculated as the ratio of the
first to the last ISI so that stronger firing frequency adaptation
is represented by a smaller adaptation ratio. In RS cells,
adaptation ratio strongly decreased with an increase of the
stimulation current, whereas in LTS cells, adaptation ratio
increased. In the majority of the IM cells, a fast initial decrease
 MONKEY PFC PYRAMIDAL CELLS 601

Table 1. Electrophysiological characteristics of pyramidal cells from different clusters

Characteristics RS_LRi RS_HRi IM LTS F Ratio P Value Fisher’s LSD

RMP, mV ⫺73.2 ⫾ 1.1 ⫺71.3 ⫾ 1.9 ⫺71.8 ⫾ 1.0 ⫺71.7 ⫾ 1.1 0.5 0.70 (1,2,3,4)
Rin, M⍀ 174 ⫾ 16 332 ⫾ 28 118 ⫾ 7 261 ⫾ 42 18.1 ⬍0.001 (3); (1); (2,4)
␶, ms 19.6 ⫾ 0.9 20.8 ⫾ 2.3 17.2 ⫾ 0.7 25.3 ⫾ 3.0 4.5 ⬍0.01 (1,2,3); (2,4)
I0, pA 106 ⫾ 8 52 ⫾ 9 120 ⫾ 7 60 ⫾ 12 12.8 ⬍0.001 (2,4); (1,3)
Sag, % 9.7 ⫾ 1.2 6.6 ⫾ 0.9 20.2 ⫾ 2.6 13.2 ⫾ 2.0 9.1 ⬍0.001 (1,2,4); (3)
RD, % 8.9 ⫾ 1.4 7.3 ⫾ 0.9 19.0 ⫾ 2.0 14.8 ⫾ 2.4 8.8 ⬍0.001 (1,2); (3; 4)
Hump, %) 11.8 ⫾ 1.9 8.6 ⫾ 2.3 31.2 ⫾ 3.7 32.3 ⫾ 4.6 14.2 ⬍0.001 (1,2); (3; 4)
TFS, ms 136 ⫾ 10 150 ⫾ 20 117 ⫾ 8 126 ⫾ 17 1.2 0.31 (1,2,3,4)
APT, mV ⫺40.4 ⫾ 0.9 ⫺40.0 ⫾ 0.9 ⫺41.4 ⫾ 0.7 ⫺44 ⫾ 0.8 3.1 ⬍0.05 (1,2,3); (3,4)
APA, mV 82 ⫾ 2 64 ⫾ 4 82 ⫾ 2 78 ⫾ 2 10.9 ⬍0.001 (2); (1,3,4)
APD, ms 0.93 ⫾ 0.02 0.88 ⫾ 0.04 0.90 ⫾ 0.03 1.12 ⫾ 0.11 3.3 ⬍0.05 (1,2,3); (4)

Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
AHP, mV 11.9 ⫾ 0.8 9.6 ⫾ 1.0 10.4 ⫾ 0.6 6.6 ⫾ 1.0 5.9 ⬍0.01 (4); (1,2,3)
fAHP, mV 7.2 ⫾ 1.5 5.5 ⫾ 1.7 5.5 ⫾ 1.1 6.0 ⫾ 1.0 0.40 0.75 (1,2,3,4)
ADP, mV 0.84 ⫾ 0.21 0.52 ⫾ 0.28 2.4 ⫾ 0.35 n/a 6.7 ⬍0.001 (1,2); (3)
mAHP, mV 5.5 ⫾ 0.7 4.6 ⫾ 0.9 7.3 ⫾ 0.6 n/a 10.1 ⬍0.001 (1,2); (3)
tAHP, ms 37 ⫾ 4 32 ⫾ 6 61 ⫾ 6 1.3 ⫾ 0.2 19.8 ⬍0.001 (4); (1,2); (3)
f1_k, Hz/pA 0.42 ⫾ 0.04 1.22 ⫾ 0.15 0.70 ⫾ 0.06 0.49 ⫾ 0.07 19.4 ⬍0.001 (1,4); (3,4); (2)
f1_threshold, Hz 1.1 ⫾ 1.2 1.0 ⫾ 5.4 16.8 ⫾ 7.7 102 ⫾ 15 34.1 ⬍0.001 (1,2,3); (4)
f2_k, Hz/pA 0.25 ⫾ 0.03 0.92 ⫾ 0.17 0.33 ⫾ 0.05 0.55 ⫾ 0.07 14.6 ⬍0.001 (1,3); (3,4); (2)
f2_threshold, Hz 1.67 ⫾ 0.65 0.52 ⫾ 2.17 ⫺1.1 ⫾ 1.1 39 ⫾ 10 23.5 ⬍0.001 (1,2,3); (4)
ISI1-2, ms 215 ⫾ 13 165 ⫾ 28 186 ⫾ 24 27 ⫾ 15 13.2 ⬍0.001 (4); (1,2,3)
FFAI, % 37 ⫾ 2 37 ⫾ 5 66 ⫾ 3 42 ⫾ 3 21.7 ⬍0.001 (1,2,4); (3)
FFAL, % 50 ⫾ 2 58 ⫾ 5 76 ⫾ 2 86 ⫾ 2 34.4 ⬍0.001 (1,2); (3); (4)
fss, Hz 16.3 ⫾ 0.7 26.6 ⫾ 2.4 17.1 ⫾ 0.9 16.5 ⫾ 1.0 13.9 ⬍0.001 (1,3,4); (2)

Values are means ⫾ SE of electrophysiological measurements in regular-spiking cells with low (RS_LRi; cluster 1; n ⫽ 27) and high input resistance
(RS_HRi; cluster 2; n ⫽ 13), intermediate-spiking cells (IM; cluster 3; n ⫽ 24), and low-threshold-spiking cells (LTS; cluster 4; n ⫽ 13). See text for definition
of electrophysiological characteristics. Following cluster analysis, Fisher’s post hoc least significant difference (LSD) test was applied to compare the means for
any pair of clusters 1– 4 with statistically distinct properties; cluster numbers grouped within parentheses for a measurement indicate no significant difference
between the indicated clusters, whereas different sets of parentheses for each measurement indicate significant differences between cells in each respective group
of clusters.

of adaptation ratio was followed by a stable adaptation ratio
(Fig. 9). When depolarizing current steps reached 150 –200 pA
above firing threshold, a similar adaptation ratio of 0.2– 0.4
was observed almost in all pyramidal cells. Thus the differ-
ences in firing pattern between pyramidal cell classes reduced
with increasing levels of depolarization.
Current-to-Frequency Transduction in Different
Electrophysiological Classes of Pyramidal Cells in
Monkey DLPFC
An essential characteristic of neuronal function is the trans-
duction of synaptic inputs into discharge. Thus we compared
how the different electrophysiological classes of pyramidal
cells transform a steady depolarizing input into AP discharge.
First, we investigated how rapidly monkey pyramidal cells
start firing in response to different intensity depolarizing cur-
rent steps. The instantaneous frequency of the first spike (f0)
(calculated as 1/latency to the 1st spike) rose linearly with the
increase of stimulation current. The slope of current vs. instan-
taneous frequency was significantly different between pyrami-
dal cell classes, being maximal in RS_HRi pyramidal cells
(0.75 ⫾ 0.01 Hz/pA), minimal in IM cells (0.30 ⫾ 0.01
Hz/pA), and intermediate in LTS (0.48 ⫾ 0.01 Hz/pA) and
RS_LRi cells (0.46 ⫾ 0.01 Hz/pA) (Fig. 10A).
Next, we examined how the discharge frequencies of differ-
ent electrophysiological classes of pyramidal cells changed
with the increase of current steps. Because pyramidal cells
exhibit firing frequency adaptation, we studied changes of
initial and steady-state frequencies by plotting them against the
amount of current injected (f/I plot). The instantaneous fre-
quency corresponding to the first ISI (f1) rose more steeply
J Neurophysiol • doi:10.1152/jn.00859.2011 • www.jn.org
with an increase of injected depolarizing current than other
instantaneous frequencies in all electrophysiological classes.
At the same time, the relationship between f1 and injected
current was cell class specific. In the majority of RS (30/40
cells) and IM cells (16/24 cells), f1 increased linearly with
current intensity. In some RS cells (10/40 cells) from both
RS_LRi and RS_HRi classes, the f1/I plot had two distinct
regions, with an initial shallow slope being followed by a
slightly steeper slope. In contrast, one-third of IM cells (8/24)
and all LTS cells exhibited a flattening of the slope (Fig. 10, C
and D). The presence of two ranges on the f1/I plot is reflected
in the linear fit parameters. The intercept of linear fit
(f1_threshold) was negative if an initial shallow slope was fol-
lowed by a steeper slope and positive in cases of flattening of
the f1/I curve. We found that f1_threshold in RS cells was close to
0 Hz (1.1 ⫾ 1.2 in RS_LRi and 1.0 ⫾ 5.4 in RS_HRi, whereas
f1_threshold was 102 ⫾ 15 Hz in the LTS cell cluster and 17 ⫾
8 Hz in the IM cell cluster). The slopes of the f1/I curve were
significantly different between pyramidal cell classes, being
maximal in RS_HRi pyramidal cells (1.22 ⫾ 0.15 Hz/pA) and
about two times smaller in other cell groups (Table 1). Thus RS
and LTS pyramidal cells were clearly separated in the
f1_threshold vs. f1_slope plot (Fig. 10F). The f2/I plots had similar
structure; however, a smaller number of cells exhibited two
distinct regions. Steady-state frequency increased linearly with
current in all cell types, and their dynamics were similar.
DISCUSSION
In this study, we examined passive and active intrinsic mem-
brane properties of layer 2/3 pyramidal cells of monkey DLPFC
and found that these neurons are electrophysiologically diverse.
602 MONKEY PFC PYRAMIDAL CELLS

Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
Fig. 4. Subthreshold membrane properties of the typical RS_HRi, RS_LRi, IM, and LTS cells in monkey DLPFC. A–D: voltage deflections to injection of a series
of hyperpolarizing and subthreshold depolarizing current steps (500 ms, 10 pA per step) of a typical neuron from each cluster. Graphs show voltage-current (V-I)
plots, where triangles indicate the maximal voltage deflection from the resting membrane potential (RMP) and circles indicate the voltage deflection at the end
of the step. Note that a time-dependent rectification in response to hyperpolarizing current steps and rebound depolarization (RD) was larger in IM and LTS cells
than in RS cells. In response to subthreshold depolarizing current steps, LTS and IM cells exhibited humplike upward voltage deflection; this was not typical
for either class of RS cells. E: bar graph summarizing the mean values of sag, hump, and RD in all 4 classes. Note the different ratio between hump and sag
in IM and LTS cells.

Our results are in contrast with data obtained from rodents, where However, electrophysiologically diverse pyramidal neurons were
almost all supragranular pyramidal cells have the characteristic found in layer 2/3 in cat and human neocortex (Chen et al. 1996;
RS firing pattern and pyramidal neurons with dissimilar firing Foehring et al. 1991; Nowak et al. 2003; Nunez et al. 1993; Tasker
patterns can be found only in infragranular layers (Agmon and et al. 1996). Thus the monkey neocortex, like human and cat
Connors 1989; Connors and Gutnick 1990; de la Pena and neocortex, appears to differ from the rodent neocortex in having a
Geijo-Barrientos 1996; Degenetais et al. 2002; Hattox and Nelson greater heterogeneity in the electrophysiological properties of
2007; Mason and Larkman 1990; Otsuka and Kawaguchi 2011). layer 2/3 pyramidal cells.
J Neurophysiol • doi:10.1152/jn.00859.2011 • www.jn.org
 MONKEY PFC PYRAMIDAL CELLS
Factors Influencing Electrophysiological Properties In Vitro
However, the direct comparison of our results with other in
vivo and in vitro studies should be done with some caution,
because many experimental factors may influence the active
and passive membrane properties of pyramidal neurons in vitro
including composition of the intracellular solution and ionic
medium, the temperature of the preparation during the electro-
physiological recordings, and the degree of dendritic tree
J Neurophysiol • doi:10.1152/jn.00859.2011 • www.jn.org
603
Fig. 5. Properties of sag, hump and RD in
monkey DLPFC pyramidal neurons. A: sag
vs. RD plot indicates a strong correlation
(r ⫽ 0.86, P ⬍ 0.001) between these param-
eters. B: sag vs. hump plot. Note that the
slope of the regression line for sag vs. hump
Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
differs between pyramidal cell classes: LTS
cells has the steepest slope. Symbols are as
indicated in A. C: voltage dependence of
humplike depolarization in LTS pyramidal
cell. The amplitude of hump increases as the
depolarizing current step is applied from
more hyperpolarized membrane potentials.
damage caused by slice preparation. For example, dendrotomy
significantly increases Rin and AP and AHP amplitudes and
hyperpolarizes the AP threshold (Bekkers and Häusser 2007).
Because some dendritic branches of pyramidal cells are
trimmed during the slice preparation, it is possible that the
electrophysiological differences between at least some cell
classes are due to differences in the degree of dendritic dam-
age. However, several lines of evidence combined indicate that
Fig. 6. Sequences of afterpotentials in different elec-
trophysiological classes of pyramidal cells in monkey
DLPFC. A–D: in the majority of pyramidal neurons,
the first action potential in a train was followed by the
same sequence of afterpotentials: a fast afterhyperpo-
larization (fAHP) was followed by a depolarizing
afterpotential (ADP) and then by a medium afterhy-
perpolarization (mAHP). The ADP was generally
more prominent in IM cells (C) than in RS cells (B);
moreover, the ADP was undetectable in some RS
neurons (A). In LTS cells, a prominent ADP led to the
next spike, which completely masked a medium-
component AHP (D). After the initial burst of spikes,
LTS cells fired single spikes with the same sequence
of afterpotentials as RS pyramidal cells. E: bar graph
summarizing the measurements of amplitude of dif-
ferent components of AHP.
604 MONKEY PFC PYRAMIDAL CELLS

Fig. 9. Changes in adaptation ratio in different electrophysiological classes of

pyramidal cells in monkey DLPFC. The changes in FFA with increasing

Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
current are cell type specific. In RS cells, adaptation ratio strongly decreased
with an increase of the stimulation current, whereas in LTS cells, adaptation
ratio increased. When depolarizing current steps reached 150 –200 pA above
firing threshold, the similar adaptation ratio of 0.2– 0.4 was observed in almost
all pyramidal cells.

Fig. 7. First interspike intervals (ISI1–2) in different electrophysiological
classes of pyramidal cells in monkey DLPFC. LTS pyramidal cells exhibit
burst firing to just suprathreshold depolarizing current pulses, and thus they
have the shortest ISI1–2 (30.5 ⫾ 19.2 ms). In other pyramidal cell groups, the
mean value of ISI1–2 is several times larger.
this factor did not confound the present classification. First, if
we assume that classes of RS_LRi and RS_HRi pyramidal cells
represent the same population of RS cells in which RS_HRi
cells are more damaged by slicing, then RS_HRi pyramidal
cells should also have larger AP and AHP amplitudes and a
more hyperpolarized APT than RS_LRi cells, as indicated by a
previous experimental study (Bekkers and Häusser 2007). In
striking contrast, RS_HRi cells have smaller AP amplitude and
similar AP threshold and amplitudes of AHP compared with
RS_LRi pyramidal cells (Table 1). Second, all pyramidal cells
included in this study had clear apical and basal dendrites, and
thus it is unlikely that differences in dendritic tree trimming

Fig. 8. Firing frequency adaptation in different electrophysiological classes of pyramidal cells in monkey DLPFC. A–D: plots of instantaneous spike frequencies
in the sequential sweeps for representative examples of pyramidal cells of 4 different electrophysiological classes. Note that with near-threshold depolarizing
current steps, both classes of RS cells (A and B) display nonadapting or weakly adapting firing pattern, whereas the LTS cells (C) exhibit strong firing frequency
adaptation; with larger depolarizing current steps, plots for RS and LTS cells look alike. In contrast, in IM cells (D), the initial spike doublet is followed by an
almost nonadapting firing with the full range of depolarizing current steps, forming the characteristic plot. E: bar graph summarizing the mean values of early
and late firing frequency adaptation (FFA) in the 4 classes. Note the different ratio between FFA_early and FFA_late in IM and LTS cells.

J Neurophysiol • doi:10.1152/jn.00859.2011 • www.jn.org

 MONKEY PFC PYRAMIDAL CELLS
during slice preparation significantly affected their electro-
physiological properties.
The ionic medium composition may determine the electrophys-
iological type of cells. Neuronal types like chattering cells or
neurons with high-frequency spike bursts have been described in
the cat’s visual, motor, and association cortical areas in vivo (Gray
and McCormick 1996; Nowak et al. 2003; Steriade et al. 1998).
However, to be detected in slices they require specific ionic
conditions, such as a 1.2 mM calcium concentration (Brumberg et
al. 2000) that appears to be closer to that of in vivo-like ionic
media at 1.2–1.4 mM (Hansen 1985). In the present study, we
used Ringer solution containing 2 mM CaCl2, the most typical
composition for in vitro studies, and thus the presence of LTS and
IM cells in layer 2/3 of monkey PFC is species specific, at least for
these in vitro conditions.
Multiple reports have demonstrated effects of the main
internal anion on the shape and the amplitude of AHP, espe-
cially on its medium and slow component (Kaczorowski et al.
2007; Zhang et al. 1994). For example, the AHP is, on average,
smaller in potassium gluconate immediately after the whole
cell configuration is obtained and remains constant for the
duration of the recording, whereas in potassium methylsulfate
J Neurophysiol • doi:10.1152/jn.00859.2011 • www.jn.org
605
Fig. 10. Current-to-frequency transduction (f/I)
in different electrophysiological classes of py-
Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
ramidal cells in monkey DLPFC. A: the aver-
aged instantaneous frequency of the first spike
(f0; calculated as 1/latency to the 1st spike)
plotted against current strength for 4 electro-
physiological classes of pyramidal cells. B–
E: representative examples of f/I plots for pyra-
midal cells from different electrophysiological
classes. The frequencies corresponding to the
first (f1), second (f2), etc., spikes increase mono-
tonically with the current strength in RS cells of
both classes (B and E) and in the IM cell (C). In
contrast, the LTS cell (D) shows an abrupt onset
of repetitive firing with increasing current for
the f1 and f2 frequencies. F: a plot of f1_threshold
vs. f1_slope demonstrates the separation of RS
and LTS pyramidal cells. fss, steady-state fre-
quency.
it is comparable to the AHP measured in the perforated-patch
configuration. Thus the direct comparison of results obtained
with sharp electrodes and patch electrodes with different com-
positions of the intracellular solution should be done with some
caution. In our experiments, potassium gluconate-based intra-
cellular solution was used, and although all registrations of
active and passive membrane properties were done during the
first 5 min after whole cell configuration was reached, we could
not exclude an underestimation of the medium component of
AHP. Because medium and slow AHP regulate firing fre-
quency and its adaptation (Faber and Sah 2007; Sah and Faber
2002), these parameters also might be affected.
Membrane properties of neurons are highly sensitive to
temperature (Ali et al. 2007; Lee et al. 2005; Thompson et al.
1985; Volgushev et al. 2000) and are altered by reductions of
only 5–10°C from normal body temperature (Thompson et al.
1985). We performed our experiments at 32°C, the most
typical temperature for such types of experiments. Thus it
should be noted that these cooling of neurons may result in an
increase in Rin, slowing of AP kinetics, and an increase in the
amplitude and duration of medium and slow AHP compared
with the normal body temperature.
606 MONKEY PFC PYRAMIDAL CELLS
Electrophysiological Classes of Layer 2/3 Pyramidal Cells in
Monkey DLPFC
We classified monkey DLPFC pyramidal cells into four
electrophysiological classes based on the results of cluster
analysis. Two classes contain cells with RS firing pattern
(RS_HRi cells and RS_LRi) and constitute 52% of the total
pyramidal cell population, another class includes LTS cells
(17%), and the last class consists of cells with an intermediate
firing pattern (31%). The two classes of RS pyramidal cells
differ in Rin, membrane time constant ␶, I0, AP amplitude, and
fss of firing (see Table 1 for a complete list of differences).
Because of the differences in intrinsic properties, RS_HRi cells
are more excitable than RS_LRi pyramidal cells, discharging
with a higher frequency than RS_LRi pyramidal at the same
level of stimulation. Thus the two classes of RS cells may play
a different functional role in DLPFC circuitry. In previous
classifications, RS cells were not separated on the basis of Rin,
with only a few exceptions to our knowledge (Kawaguchi
1993; Schwindt et al. 1997). Usually, RS pyramidal cells were
divided into two or three subgroups based primarily on the
degree of changes in AP threshold and spike frequency adap-
tation across a sustained train of APs (Agmon and Connors
1992; Chang and Luebke 2007; Degenetais et al. 2002). In
contrast to these reports, we usually did not observe the
changes in AP threshold or amplitude across the train of APs
in monkey DLPFC RS cells in a range of applied depolariza-
tion current. Also, in our sample we did not detect cells with
properties known as RS fast-adapting pyramidal cells, which
fire a brief train of 2–10 APs followed by a depolarizing
plateau (Chang and Luebke 2007; Degenetais et al. 2002;
Nunez et al. 1993; Otsuka and Kawaguchi 2008). The monkey
DLPFC RS cells described in this study are most similar to
neurons previously reported in rodents as RS1 (Agmon and
Connors 1992), RS (Kawaguchi 1993; Mason and Larkman
1990; Yang et al. 1996), or slow-adapting RS1 cells (Degen-
etais et al. 2002); in cats as RS slow-adapting (Nunez et al.
1993) or RS cells (Chen et al. 1996; Nowak et al. 2003); in
monkey as non-LTS RS1 cells (Chang and Luebke 2007); and
in humans as RS (Foehring et al. 1991) and non-LTS cells
(Tasker et al. 1996). Similar to RS cells from other species, RS
cells in monkey DLPFC had little or no anomalous inward
rectification detected in either the hyperpolarized or the depo-
larized membrane voltage range.
Here we report that at least one-third of pyramidal cells in
layer 2/3 of monkey DLPFC exhibit LTS and IM firing
patterns. In the LTS cells, the burst of APs was evoked at I0,
whereas a larger current is required to evoke bursts in IM
pyramidal cells. The bursts were typically two to five spikes
long; two-spike bursts were most common in IM cells, whereas
LTS cells commonly fired bursts with three to five spikes. As
mentioned above, cells with burst firing are usually found only
in deep layers in the rodent neocortex (de la Pena and Geijo-
Barrientos 1996; Larkum et al. 2007; Mason and Larkman
1990; Schwindt et al. 1997). For example, in rodent medial
PFC, LTS cells constitute up to 64 – 68% in layers 5 and 6 and
⬍5% in layer 2/3 (de la Pena and Geijo-Barrientos 1996;
Otsuka and Kawaguchi 2011; Yang et al. 1996). However, it
was reported that two initial high-frequency spikes (“dou-
blets”) can be evoked in cells in all cortical layers (Agmon and
Connors 1992; Connors 1984; McCormick et al. 1985). It is
J Neurophysiol • doi:10.1152/jn.00859.2011 • www.jn.org
also worth noting that LTS cells in human neocortex are more
similar to monkey IM cells than to monkey LTS cells, because
at depolarized potentials (more positive than ⫺69 mV), LTS
neurons in human cortex respond with RS firing, whereas at
more hyperpolarized potentials they respond with two- or,
rarely, three-spike bursts that ride on a hump and are followed
by rhythmic spiking (Foehring et al. 1991; Tasker et al. 1996).
The LTS firing pattern is believed to be generated by
low-threshold calcium channels (LTCC) (de la Pena and Geijo-
Barrientos 1996; Yang et al. 1996). In the present study, we did
not directly investigate the ionic basis of the LTS firing pattern
in monkey PFC neurons; however, some observations suggest
Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
the presence of LTCCs in monkey LTS and IM pyramidal
cells. For example, we observed that the humplike depolariza-
tion in LTS and IM neurons was much more intense if the
depolarizing current was applied at a more hyperpolarized
membrane potential. In contrast, RS pyramidal cells typically
did not exhibit hump at any membrane potential. Another sign
of LTCC contribution is the presence of postspike ADP (Yang
et al. 1996). Although the ionic basis for the postspike ADPs
differs widely in various central neurons (Higashi et al. 1993),
in rat PFC neurons, Ca2⫹ entry plays a critical role in electro-
genesis of ADPs (Yang et al. 1996). IM cells possessed a large
postspike ADP, in LTS cells the bursting complex also was
derived from a ADP that followed the first AP, whereas ADP
was less prominent or completely absent in RS pyramidal cells.
Functional Significance of the Findings
The DLPFC plays a key role in complex cognitive functions,
for instance, working memory. In fact, it is generally assumed
that the sustained firing of DLPFC neurons during the delay
period of working memory tasks represents the active holding
of relevant information required for correct task performance
(Goldman-Rakic 1995). The ability of DLPFC pyramidal neu-
rons to sustain delay period firing during working memory
tasks may be influenced by the intrinsic membrane properties
of these cells. For example, strong firing frequency adaptation
during sustained excitatory input opposes the production of
persistent delay-related firing via recurrent excitation (Carter
and Wang 2007; Pinto and Ermentrout 2001). Consequently,
strong spike frequency adaptation may decrease the efficacy of
storage of information in working memory, as suggested by
computational modeling studies (Carter and Wang 2007). In-
terestingly, in the present study we did not observe fast-
adapting RS cells (Chang and Luebke 2007; Degenetais et al.
2002; Nunez et al. 1993; Otsuka and Kawaguchi 2008). More-
over, no firing frequency adaptation was observed in RS cells
if low-intensity depolarizing current was applied (Fig. 9).
Under larger depolarizing current steps, the firing frequency
remained relatively stable after the short period of rate adap-
tation: LTS and RS_HRi cells adapted within the first three to
six spikes (the first 100 –200 ms after the onset of the current
pulse), and IM and RS_LRi cells did not change their firing
rates after the first ISI (Fig. 8, A, B, and E). These data suggest
that among the different pyramidal cell classes described in this
study, RS cells appear to have the most appropriate electro-
physiological properties for maintaining persistent delay-re-
lated firing.
During working memory tasks, some primate DLPFC neu-
rons display brief cue- or response-related activity instead of
 MONKEY PFC PYRAMIDAL CELLS
sustained delay period-related firing (Goldman-Rakic 1996).
An important question is, therefore, whether differences in
biophysical membrane properties, which strongly modulate the
response to brief or sustained excitatory input, contribute to the
different patterns of task-related firing of primate DLPFC
neurons. However, many, if not most, individual DLPFC
neurons respond in more than one phase of the trial, i.e., during
the cue, delay, and/or response periods (Goldman-Rakic 1996).
Thus the heterogeneity of task-related activity of most primate
DLPFC neurons cannot be determined by biophysical proper-
ties alone and probably requires the combined effects of
intrinsic membrane properties, specific morphological proper-
ties, and functional connectivity. For example, the composite
firing profile of DLPFC cells with activity in more than one
phase of the task may be due to inputs from neurons whose
activation is simpler and related to only one phase (Goldman-
Rakic 1996).
Neuronal activity in primate DLPFC is crucial for cognitive
control, a cognitive mechanism allowing coordination of lower
level sensory and motor operations and memory to achieve a
specific goal (Miller 2000; Miller and Cohen 2001). Cognitive
control must implicate complex forms of information process-
ing in DLPFC circuits, which may require specific forms of infor-
mation transmission via DLPFC neuron activity. Interestingly,
intrinsic bursting neurons may play unique roles in information
transmission combined with specific forms of short-term synaptic
plasticity. For instance, bursts may enhance transmission of in-
formation to downstream neurons (Lisman 1997), enable coding
of changes in network firing rate (Abbott et al. 1997), or enhance
recruitment of specific interneuron subtypes (Berger et al.
2010). Importantly, whereas LTS cells had bursting properties
irrespective of the depolarizing stimulus strength, IM cells
displayed two modes of response, regular spiking and bursting
firing, depending on the strength of stimulation and membrane
potential value. Thus the proportion of monkey DLPFC neu-
rons with bursting activity may vary according to the overall
level of network activity.
Differential excitability of pyramidal cell classes and the
diversity of current-to-frequency transduction properties may
play an important role in cortical network functioning. For
example, different cell types would be involved in firing with
different strengths of synaptic stimulation: LTS and RS_HRi
pyramidal cells need smaller depolarizing current to fire, and
moreover, because they have a longer membrane time constant,
they may be more efficient at summation of excitatory synaptic
potentials. In contrast, IM and RS_LRi will be activated with
stronger stimuli. RS cells are well suited to perform a linear
transduction of synaptic inputs to spike output over a wide
range of currents with different gain; the RS_LRi cells have a
significantly smaller gain (f/I curve slope) than the RS_HRi
cells and would thus fire fewer spikes in response to the same
synaptic stimulation. It is worth noting that the cell excitability
and the transduction gain are regulated by many neuromodu-
lators, which have multiple effects on intrinsic membrane
properties and synaptic neurotransmission (Saar and Barkai
2009; Seamans and Yang 2004). For example, dopamine neu-
romodulation strongly affects the excitability of primate
DLPFC pyramidal neurons (Henze et al. 2000) and their firing
in vivo during working memory tasks (Sawaguchi 2001; Wang
et al. 2004; Williams and Goldman-Rakic 1995). Moreover,
extracellular dopamine levels in monkey DLPFC rise at the
J Neurophysiol • doi:10.1152/jn.00859.2011 • www.jn.org
607
beginning of a working memory task and stay elevated across
many trials (Watanabe et al. 1997). A number of studies in both
rat and primate PFC have confirmed that, at least under certain
conditions, dopamine via D1 receptors increases the excitabil-
ity of PFC neurons in vitro and in vivo (Seamans and Yang
2004) and thus changes their output gain. Our present results
suggest that heterogeneity in the intrinsic firing properties of
pyramidal cells may contribute to information processing in
superficial layers of the DLPFC during cognitive function.
Further studies are necessary to fully understand the complex
interactions between biophysical properties of individual
DLPFC neurons and other multiple factors that ultimately
determine their patterns of activity in vivo during cognitive
Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
tasks.
GRANTS
This work was supported by National Institute of Mental Health Grants
MH043784 and MH084053. A. Zaitsev was also supported by Russian Foun-
dation for Basic Research Grant 11-04-00912
and by the Program for Basic
Research of the Presidium of the Russian Academy of Sciences (Molecular and
Cell Biology).
DISCLOSURES
D. A. Lewis currently receives investigator-initiated research support from
the BMS Foundation, Bristol-Myers Squibb, Curridium Ltd., and Pfizer and in
2010 –2012 served as a consultant in the areas of target identification and
validation and new compound development to Bristol-Myers Squibb.
AUTHOR CONTRIBUTIONS
Author contributions: A.V.Z., N.V.P., G.G.-B., and D.A.L. conception and
design of research; A.V.Z., N.V.P., G.G.-B., and D.A.L. performed experi-
ments; A.V.Z. analyzed data; A.V.Z., N.V.P., G.G.-B., and D.A.L. interpreted
results of experiments; A.V.Z. prepared figures; A.V.Z. and G.G.-B. drafted
manuscript; A.V.Z., N.V.P., G.G.-B., and D.A.L. edited and revised manu-
script; A.V.Z., N.V.P., G.G.-B., and D.A.L. approved final version of manu-
script.
REFERENCES
Abbott LF, Varela JA, Sen K, Nelson SB. Synaptic depression and cortical
gain control. Science 275: 220 –224, 1997.
Agmon A, Connors BW. Correlation between intrinsic firing patterns and
thalamocortical synaptic responses of neurons in mouse barrel cortex. J
Neurosci 12: 319 –329, 1992.
Agmon A, Connors BW. Repetitive burst-firing neurons in the deep layers of
mouse somatosensory cortex. Neurosci Lett 99: 137–141, 1989.
Ali AB, Bannister AP, Thomson AM. Robust correlations between action
potential duration and the properties of synaptic connections in layer 4
interneurones in neocortical slices from juvenile rats and adult rat and cat.
J Physiol 580: 149 –169, 2007.
Artchakov D, Tikhonravov D, Ma Y, Neuvonen T, Linnankoski I, Carlson
S. Distracters impair and create working memory-related neuronal activity
in the prefrontal cortex. Cereb Cortex 19: 2680 –2689, 2009.
Baddeley A. Working memory: looking back and looking forward. Nat Rev
Neurosci 4: 829 – 839, 2003.
Bekkers JM, Häusser M. Targeted dendrotomy reveals active and passive
contributions of the dendritic tree to synaptic integration and neuronal
output. Proc Natl Acad Sci USA 104: 11447–11452, 2007.
Berger TK, Silberberg G, Perin R, Markram H. Brief bursts self-inhibit and
correlate the pyramidal network. PLoS Biol 8: pii: e1000473, 2010.
Brumberg JC, Nowak LG, McCormick DA. Ionic mechanisms underlying
repetitive high-frequency burst firing in supragranular cortical neurons. J
Neurosci 20: 4829 – 4843, 2000.
Carter E, Wang XJ. Cannabinoid-mediated disinhibition and working mem-
ory: dynamical interplay of multiple feedback mechanisms in a continuous
attractor model of prefrontal cortex. Cereb Cortex 17, Suppl 1: i16 –i26,
2007.
608 MONKEY PFC PYRAMIDAL CELLS
Chang YM, Luebke JI. Electrophysiological diversity of layer 5 pyramidal
cells in the prefrontal cortex of the rhesus monkey: in vitro slice studies. J
Neurophysiol 98: 2622–2632, 2007.
Chang YM, Rosene DL, Killiany RJ, Mangiamele LA, Luebke JI. In-
creased action potential firing rates of layer 2/3 pyramidal cells in the
prefrontal cortex are significantly related to cognitive performance in aged
monkeys. Cereb Cortex 15: 409 – 418, 2005.
Chen W, Zhang JJ, Hu GY, Wu CP. Electrophysiological and morpholog-
ical properties of pyramidal and nonpyramidal neurons in the cat motor
cortex in vitro. Neuroscience 73: 39 –55, 1996.
Connors BW. Initiation of synchronized neuronal bursting in neocortex.
Nature 310: 685– 687, 1984.
Connors BW, Gutnick MJ. Intrinsic firing patterns of diverse neocortical
neurons. Trends Neurosci 13: 99 –104, 1990.
Curtis CE, Lee D. Beyond working memory: the role of persistent activity in
decision making. Trends Cogn Sci 14: 216 –222, 2010.
de la Pena E, Geijo-Barrientos E. Laminar localization, morphology, and
physiological properties of pyramidal neurons that have the low-threshold
calcium current in the guinea-pig medial frontal cortex. J Neurosci 16:
5301–5311, 1996.
Degenetais E, Thierry AM, Glowinski J, Gioanni Y. Electrophysiological
properties of pyramidal neurons in the rat prefrontal cortex: an in vivo
intracellular recording study. Cereb Cortex 12: 1–16, 2002.
Elston GN. Cortex, cognition and the cell: new insights into the pyramidal
neuron and prefrontal function. Cereb Cortex 13: 1124 –1138, 2003.
Elston GN, Benavides-Piccione R, DeFelipe J. The pyramidal cell in cog-
nition: a comparative study in human and monkey. J Neurosci 21: RC163,
2001.
Faber ES, Sah P. Functions of SK channels in central neurons. Clin Exp
Pharmacol Physiol 34: 1077–1083, 2007.
Foehring RC, Lorenzon NM, Herron P, Wilson CJ. Correlation of physi-
ologically and morphologically identified neuronal types in human associ-
ation cortex in vitro. J Neurophysiol 66: 1825–1837, 1991.
Fuster JM. Network memory. Trends Neurosci 20: 451– 459, 1997.
Goldman-Rakic PS. Cellular basis of working memory. Neuron 14: 477– 485,
1995.
Goldman-Rakic PS. Regional and cellular fractionation of working memory.
Proc Natl Acad Sci USA 93: 13473–13480, 1996.
Gonzalez-Burgos G, Barrionuevo G, Lewis DA. Horizontal synaptic con-
nections in monkey prefrontal cortex: an in vitro electrophysiological study.
Cereb Cortex 10: 82–92, 2000.
Gonzalez-Burgos G, Krimer LS, Povysheva NV, Barrionuevo G, Lewis
DA. Functional properties of fast spiking interneurons and their synaptic
connections with pyramidal cells in primate dorsolateral prefrontal cortex. J
Neurophysiol 93: 942–953, 2005a.
Gonzalez-Burgos G, Krimer LS, Urban NN, Barrionuevo G, Lewis DA.
Synaptic efficacy during repetitive activation of excitatory inputs in primate
dorsolateral prefrontal cortex. Cereb Cortex 14: 530 –542, 2004.
Gonzalez-Burgos G, Kroener S, Seamans JK, Lewis DA, Barrionuevo G.
Dopaminergic modulation of short-term synaptic plasticity in fast-spiking
interneurons of primate dorsolateral prefrontal cortex. J Neurophysiol 94:
4168 – 4177, 2005b.
Gonzalez-Burgos G, Kroener S, Zaitsev AV, Povysheva NV, Krimer LS,
Barrionuevo G, Lewis DA. Functional maturation of excitatory synapses in
layer 3 pyramidal neurons during postnatal development of the primate
prefrontal cortex. Cereb Cortex 18: 626 – 637, 2008.
Gonzalez-Burgos G, Rotaru DC, Zaitsev AV, Povysheva NV, Lewis DA.
GABA transporter GAT1 prevents spillover at proximal and distal GABA
synapses onto primate prefrontal cortex neurons. J Neurophysiol 101:
533–547, 2009.
Gray CM, McCormick DA. Chattering cells: superficial pyramidal neurons
contributing to the generation of synchronous oscillations in the visual
cortex. Science 274: 109 –113, 1996.
Hansen AJ. Effect of anoxia on ion distribution in the brain. Physiol Rev 65:
101–148, 1985.
Hattox AM, Nelson SB. Layer V neurons in mouse cortex projecting to
different targets have distinct physiological properties. J Neurophysiol 98:
3330 –3340, 2007.
Henze DA, Gonzalez-Burgos GR, Urban NN, Lewis DA, Barrionuevo G.
Dopamine increases excitability of pyramidal neurons in primate prefrontal
cortex. J Neurophysiol 84: 2799 –2809, 2000.
Higashi H, Tanaka E, Inokuchi H, Nishi S. Ionic mechanisms underlying the
depolarizing and hyperpolarizing afterpotentials of single spike in guinea-
pig cingulate cortical neurons. Neuroscience 55: 129 –138, 1993.
J Neurophysiol • doi:10.1152/jn.00859.2011 • www.jn.org
Hikosaka O, Takikawa Y, Kawagoe R. Role of the basal ganglia in the
control of purposive saccadic eye movements. Physiol Rev 80: 953–978,
2000.
Jacobs B, Schall M, Prather M, Kapler E, Driscoll L, Baca S, Jacobs J,
Ford K, Wainwright M, Treml M. Regional dendritic and spine variation
in human cerebral cortex: a quantitative Golgi study. Cereb Cortex 11:
558 –571, 2001.
Johnson RA, Wichern DW. Applied Multivariate Statistical Analysis. Upper
Saddle River, NJ: Prentice Hall, 1998.
Kaczorowski CC, Disterhoft J, Spruston N. Stability and plasticity of
intrinsic membrane properties in hippocampal CA1 pyramidal neurons:
effects of internal anions. J Physiol 578: 799 – 818, 2007.
Kawaguchi Y. Groupings of nonpyramidal and pyramidal cells with specific
physiological and morphological characteristics in rat frontal cortex. J
Neurophysiol 69: 416 – 431, 1993.
Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
Krimer LS, Zaitsev AV, Czanner G, Kroner S, Gonzalez-Burgos G,
Povysheva NV, Iyengar S, Barrionuevo G, Lewis DA. Cluster analysis-
based physiological classification and morphological properties of inhibitory
neurons in layers 2–3 of monkey dorsolateral prefrontal cortex. J Neuro-
physiol 94: 3009 –3022, 2005.
Kritzer MF, Goldman-Rakic PS. Intrinsic circuit organization of the major
layers and sublayers of the dorsolateral prefrontal cortex in the rhesus
monkey. J Comp Neurol 359: 131–143, 1995.
Larkman A, Mason A. Correlations between morphology and electrophysi-
ology of pyramidal neurons in slices of rat visual cortex. I. Establishment of
cell classes. J Neurosci 10: 1407–1414, 1990.
Larkum ME, Waters J, Sakmann B, Helmchen F. Dendritic spikes in apical
dendrites of neocortical layer 2/3 pyramidal neurons. J Neurosci 27: 8999 –
9008, 2007.
Lee JC, Callaway JC, Foehring RC. Effects of temperature on calcium
transients and Ca2⫹-dependent afterhyperpolarizations in neocortical pyra-
midal neurons. J Neurophysiol 93: 2012–2020, 2005.
Lisman JE. Bursts as a unit of neural information: making unreliable synapses
reliable. Trends Neurosci 20: 38 – 43, 1997.
Luebke J, Barbas H, Peters A. Effects of normal aging on prefrontal area 46
in the rhesus monkey. Brain Res Rev 62: 212–232, 2010.
Luebke JI, Amatrudo JM. Age-related increase of sIAHP in prefrontal
pyramidal cells of monkeys: relationship to cognition. Neurobiol Aging 33:
1085–1095, 2012.
Maravall M, Stern EA, Svoboda K. Development of intrinsic properties and
excitability of layer 2/3 pyramidal neurons during a critical period for
sensory maps in rat barrel cortex. J Neurophysiol 92: 144 –156, 2004.
Mason A, Larkman A. Correlations between morphology and electrophysi-
ology of pyramidal neurons in slices of rat visual cortex. II. Electrophysi-
ology. J Neurosci 10: 1415–1428, 1990.
McCormick DA, Connors BW, Lighthall JW, Prince DA. Comparative
electrophysiology of pyramidal and sparsely spiny stellate neurons of the
neocortex. J Neurophysiol 54: 782– 806, 1985.
Melchitzky DS, Gonzalez-Burgos G, Barrionuevo G, Lewis DA. Synaptic
targets of the intrinsic axon collaterals of supragranular pyramidal neurons
in monkey prefrontal cortex. J Comp Neurol 430: 209 –221, 2001.
Melchitzky DS, Sesack SR, Pucak ML, Lewis DA. Synaptic targets of
pyramidal neurons providing intrinsic horizontal connections in monkey
prefrontal cortex. J Comp Neurol 390: 211–224, 1998.
Miller EK. The prefrontal cortex and cognitive control. Nat Rev Neurosci 1:
59 – 65, 2000.
Miller EK, Cohen JD. An integrative theory of prefrontal cortex function.
Annu Rev Neurosci 24: 167–202, 2001.
Miller EK, Erickson CA, Desimone R. Neural mechanisms of visual working
memory in prefrontal cortex of the macaque. J Neurosci 16: 5154 –5167,
1996.
Nowak LG, Azouz R, Sanchez-Vives MV, Gray CM, McCormick DA.
Electrophysiological classes of cat primary visual cortical neurons in vivo as
revealed by quantitative analyses. J Neurophysiol 89: 1541–1566, 2003.
Nunez A, Amzica F, Steriade M. Electrophysiology of cat association cortical
cells in vivo: intrinsic properties and synaptic responses. J Neurophysiol 70:
418 – 430, 1993.
Otsuka T, Kawaguchi Y. Cell diversity and connection specificity between
callosal projection neurons in the frontal cortex. J Neurosci 31: 3862–3870,
2011.
Otsuka T, Kawaguchi Y. Firing-pattern-dependent specificity of cortical
excitatory feed-forward subnetworks. J Neurosci 28: 11186 –11195, 2008.
Pape HC. Queer current and pacemaker: the hyperpolarization-activated
cation current in neurons. Annu Rev Physiol 58: 299 –327, 1996.
 MONKEY PFC PYRAMIDAL CELLS
Pinto D, Ermentrout G. Spatially structured activity in synaptically coupled
neuronal networks: I. Traveling fronts and pulses. SIAM J Appl Math 62:
206, 2001.
Povysheva NV, Gonzalez-Burgos G, Zaitsev AV, Kroner S, Barrionuevo
G, Lewis DA, Krimer LS. Properties of excitatory synaptic responses in
fast-spiking interneurons and pyramidal cells from monkey and rat prefron-
tal cortex. Cereb Cortex 16: 541–552, 2006.
Povysheva NV, Zaitsev AV, Kroner S, Krimer OA, Rotaru DC, Gonzalez-
Burgos G, Lewis DA, Krimer LS. Electrophysiological differences be-
tween neurogliaform cells from monkey and rat prefrontal cortex. J Neuro-
physiol 97: 1030 –1039, 2007.
Povysheva NV, Zaitsev AV, Rotaru DC, Gonzalez-Burgos G, Lewis DA,
Krimer LS. Parvalbumin-positive basket interneurons in monkey and rat
prefrontal cortex. J Neurophysiol 100: 2348 –2360, 2008.
Pucak ML, Levitt JB, Lund JS, Lewis DA. Patterns of intrinsic and
associational circuitry in monkey prefrontal cortex. J Comp Neurol 376:
614 – 630, 1996.
Robinson RB. Hyperpolarization-activated cation currents: from molecules to
physiological function. Annu Rev Physiol 65: 453– 480, 2003.
Saar D, Barkai E. Long-lasting maintenance of learning-induced enhanced
neuronal excitability: mechanisms and functional significance. Mol Neuro-
biol 39: 171–177, 2009.
Sah P, Faber ES. Channels underlying neuronal calcium-activated potassium
currents. Prog Neurobiol 66: 345–353, 2002.
Sawaguchi T. The effects of dopamine and its antagonists on directional
delay-period activity of prefrontal neurons in monkeys during an oculomo-
tor delayed-response task. Neurosci Res 41: 115–128, 2001.
Schwindt P, O’Brien JA, Crill W. Quantitative analysis of firing properties
of pyramidal neurons from layer 5 of rat sensorimotor cortex. J Neuro-
physiol 77: 2484 –2498, 1997.
Seamans JK, Yang CR. The principal features and mechanisms of dopamine
modulation in the prefrontal cortex. Prog Neurobiol 74: 1–58, 2004.
Selemon LD, Goldman-Rakic PS. Common cortical and subcortical targets
of the dorsolateral prefrontal and posterior parietal cortices in the rhesus
monkey: evidence for a distributed neural network subserving spatially
guided behavior. J Neurosci 8: 4049 – 4068, 1988.
Steriade M, Timofeev I, Durmuller N, Grenier F. Dynamic properties of
corticothalamic neurons and local cortical interneurons generating fast
rhythmic (30 – 40 Hz) spike bursts. J Neurophysiol 79: 483– 490, 1998.
Tasker JG, Hoffman NW, Kim YI, Fisher RS, Peacock WJ, Dudek FE.
Electrical properties of neocortical neurons in slices from children with
intractable epilepsy. J Neurophysiol 75: 931–939, 1996.
J Neurophysiol • doi:10.1152/jn.00859.2011 • www.jn.org
609
Thompson SM, Masukawa LM, Prince DA. Temperature dependence of
intrinsic membrane properties and synaptic potentials in hippocampal CA1
neurons in vitro. J Neurosci 5: 817– 824, 1985.
Urban NN, Gonzalez-Burgos G, Henze DA, Lewis DA, Barrionuevo G.
Selective reduction by dopamine of excitatory synaptic inputs to pyramidal
neurons in primate prefrontal cortex. J Physiol 539: 707–712, 2002.
Volgushev M, Vidyasagar TR, Chistiakova M, Yousef T, Eysel UT.
Membrane properties and spike generation in rat visual cortical cells during
reversible cooling. J Physiol 522: 59 –76, 2000.
Wang M, Gamo NJ, Yang Y, Jin LE, Wang XJ, Laubach M, Mazer JA,
Lee D, Arnsten AF. Neuronal basis of age-related working memory
decline. Nature 476: 210 –213, 2011.
Wang M, Vijayraghavan S, Goldman-Rakic PS. Selective D2 receptor
actions on the functional circuitry of working memory. Science 303: 853–
856, 2004.
Downloaded from http://jn.physiology.org/ at St Josephs Hosp & Medical Ctr on April 10, 2013
Wang XJ. Synaptic reverberation underlying mnemonic persistent activity.
Trends Neurosci 24: 455– 463, 2001.
Ward JH. Hierarchical grouping to optimize an objective function. J Am Stat
Assoc 58: 236 –244, 1963.
Watanabe M, Kodama T, Hikosaka K. Increase of extracellular dopamine in
primate prefrontal cortex during a working memory task. J Neurophysiol 78:
2795–2798, 1997.
Watanabe Y, Funahashi S. Neuronal activity throughout the primate medi-
odorsal nucleus of the thalamus during oculomotor delayed-responses. I.
Cue-, delay-, and response-period activity. J Neurophysiol 92: 1738 –1755,
2004.
Williams GV, Goldman-Rakic PS. Modulation of memory fields by dopa-
mine D1 receptors in prefrontal cortex. Nature 376: 572–575, 1995.
Wood JN, Grafman J. Human prefrontal cortex: processing and representa-
tional perspectives. Nat Rev Neurosci 4: 139 –147, 2003.
Yang CR, Seamans JK, Gorelova N. Electrophysiological and morphological
properties of layers V-VI principal pyramidal cells in rat prefrontal cortex in
vitro. J Neurosci 16: 1904 –1921, 1996.
Zaitsev AV, Gonzalez-Burgos G, Povysheva NV, Kroner S, Lewis DA,
Krimer LS. Localization of calcium-binding proteins in physiologically and
morphologically characterized interneurons of monkey dorsolateral prefron-
tal cortex. Cereb Cortex 15: 1178 –1186, 2005.
Zaitsev AV, Povysheva NV, Gonzalez-Burgos G, Rotaru D, Fish KN,
Krimer LS, Lewis DA. Interneuron diversity in layers 2–3 of monkey
prefrontal cortex. Cereb Cortex 19: 1597–1615, 2009.
Zhang L, Weiner JL, Valiante TA, Velumian AA, Watson PL, Jahromi
SS, Schertzer S, Pennefather P, Carlen PL. Whole-cell recording of the
Ca2⫹-dependent slow afterhyperpolarization in hippocampal neurones: ef-
fects of internally applied anions. Pflügers Arch 426: 247–253, 1994.