Relationships among Unit Discharge Rate, Pattern, and Phasic Arousal in the Medial Geniculate ...

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EXPERIMENTAL KEUROLOGl- 35, 337-357 (1972)
Relationships among Unit Discharge Rate, Pattern,
and Phasic Arousal in the Medial Geniculate
Nucleus of the Waking Cat
THOMAS J. IMIG, NORMAX AI. \VEINRERGER, AKD IRWIS S.\?~ESTENBERG'
Dcpavtmmf of Psychobiology, UlliTrersify of California. Iwirw, Crrlifovnia 92664
Recek~cd Jnuuary 6, lQi2
The present experiment was designed to investigate some of the labile discharge
characteristics of single units in the medial geniculate body (MGB) of the locally
anesthetized, paralyzed cat. In particular, the following problems were addressed :
To what extent do MGB cells exhibit stationary discharge rates; what are the
effects of phasic arousal (produced by infrequent shock to the paw) upon the
discharge rate of units; and how does the pattern of unit discharge relate to its
rate? Only 4 out of 20 units showed stationary discharge rates throughout the
entire time they were studied. Furthermore approximately 50% of the cells
showed changes in discharge rate prior to the occurrence of paw shock indicating
that nonstationary discharge rate cannot be attributed entirely to shock. When
discharge rates preceding shock xvere compared with discharge rates following
shock, a change in rate was seen in about 50% of the units of which the majority
showed increases in rate. The discharge rates of single units in this prepara-
tion may apparently change both spontaneously and in response to paw shock.
W-hen the poststimulus time histograms (PSTH) were analyzed with respect to
the over-all discharge rate of the cell, several striking features became apparent.
If the rate was stationary, the PSTH generally showed no systematic variations
in shape. If the rate was nonstationary, however, the PSTH systematically dis-
played certain properties. The initial short latency peak (< 20 msec) showed no
changes corresponding to changes in rate. In contrast to the first peak, the longer
latency peaks (SO-300 msec) varied systematically with the discharge rate. These
observations were verified statistically. The results are discussed with reference
to the role of sensory systems in hehavioral plasticity.
introduction
Approaches to the function of the central auditory system have generally
taken one of two paths: analysis of single unit discharges as a function of
systematic variation of stimulus parameters using a steady-state prepara-
tion ; or analysis of evoked slow-wave potentials (EP) using a constant
l Supported by U.S.P.H.S. Grant MH 11250. The authors thank Mrs. C. Wells for
technical assistance and Mrs. C. Johnson and Miss S. Harris for secretarial services.
Dr. Imig holds Predoctoral Fellowship MH 49757. Dr. Westenberg’s studies were
supported by U.S.P.H.S. Training Center Grant RIH 11095.
337
Q 1972 by Academic Press, Ioc
338 IMIG, WEINBERGER, AND WESTENBERG
stimulus but a preparation which is likely to vary in arousal level. The
first approach characterizes sensory neurophysiology and it has revealed
the intricate transformations which appear at various levels of the mam-
malian auditory system, e.g., binaural interaction at the superior olivary
complex (3)) selective responsiveness to frequency-modulated tones at the
auditory cortex (19). The second approach is used by investigators seek-
ing to understand the dynamic properties of the auditory system as a func-
tion of behavioral or psychological variables such as habituation (lS), dis-
traction ( 10 j, learning (8, 12), sleep-waking (7, 17), and arousal
produced by reticular formation stimulation ( 16 j . As these approaches dif-
fer in type of evoked activity, degree of stimulus manipulation, and varia-
tion in organismic state, the findings from one are not easily related to the
other. In particular, there is a paucity of information regarding the dy-
namic properties of single cells. One attempt to bridge this gap involves
the analysis of multiple-unit or “fast” activity, rather than EP, in behav-
iorally relevant situations, e.g., habituation (9, ll), shock-pairing (4), and
conditioning (6).
The present experiment is an initial attempt to analyze both stable and
dynamic aspects of single unit discharges in the medial geniculate body
(MGB) of the waking cat. In this study the dynamics of unit firing are
approached by producing a phasic increase in arousal level by the infre-
quent administration of paw shock. We have been concerned with the fol-
lowing issues: (a) To what extent do MGB cells exhibit rate stationarity
as a function of time ? (b) What are the effects of phasic arousal upon the
rate of unit discharges? (c) How does the pattern of unit discharge relate
to its rate? (d) What are the effects of phasic arousal upon discharge pat-
terns? A subsequent study is concerned with some of these issues during
click-shock pairings (unpubIished data).
Methods
Subjects and Swgical Procedure. The present esperiments were carried
out on 23 cats, eight of which yielded the 20 units which were held long
enough to include in this study. The cats. weighing 2.5-3.5 kg, to be in-
cluded in the experiment, were required to respond with an orienting re-
sponse to a sudden sound, and to be free of middle ear infection (as evi-
denced by normal light reflection from the tympanic membrane during
otoscopic examination). All surgical and experimental procedures were
carried out with the cat enclosed in an acoustically dampened room (I;%C
1202). The animals were initially anesthetized with intravenous sodium
thiamylal (Surital, Parke-Davis). a short lasting barbituate anesthetic ; ad-
ditional doses were given as often as necessary to maintain a surgical level
of anesthesia. To prevent respiratory congestion, the cats in the latter half
339
MEDIAL GENICULATE NUCLEUS
of the study were given atropine methylsulfate (Burrough Wellcome ; 1
mg, ip). The left femoral vein was cannulated to allow subsequent infusion
of Surital and gallamine triethiodide (Flaxedil, Davis-Geck) and an endo-
tracheal tube with an inflatable cuff was inserted into the trachea with the
aid of a laryngoscope. The animal was mounted in a stereotaxic frame
with blunt hollow ear bars through which acoustic stimuli were presented.
Body temperature was maiutainetl at approximately 37 C by a circulating
water pad and monitored with a rectal thermometer. The cranium was ex-
posed with a midsagittal incision and the overlying muscle and connective
tissue were deflected to the sides. Trephine holes were placed to allow ac-
cess to the MGB and mesencephalic reticular formation (MRF) , and the
dura mater was cut and retracted. Two screws were placed in the skull, one
overlying the middle suprasylvian gyrus to monitor the EEG and the other
over the frontal sinus to serve as a reference electrode. The cochlear AP
and CM were monitored in some animals from a wire placed on the round
window. Shock electrodes were attached to the forepaw contralateral to the
side of the brain in which the MGB and MRF electrodes were placed and
the maintenance of general anesthesia was discontinued. All wound edges
and pressure points were heavily infiltrated with Zyljectin (Abbott), a
long lasting local anesthetic. When the animal began to show spontaneous
movements, the endotracheal tube was connected to a respirator and Flax-
edil (2 ml, iv) was injected. Paralysis was maintained throughout the ex-
periment with gravity infusion of Flaxedil diluted to 10% with dextrose-
saline solution.
Stimulation and Recording. An Epoxilite insulated stainless steel insect
pin with a resistance of 20-50Kohms served to record multiple unit activ-
ity in the MRF and a glass-coated tungsten electrode with a tip size 1-5,~
and an impedance of 3-10 Megohms at 1KHz was used to record single
neurons in the MGB. Tektronix model 122 preamplifiers provided the first
stage of amplification of the bioelectric activity. The two channels amplify-
ing MGB and MRF activity had filter settings of SO-10 KHz; the third
amplifier bandpass was 0.8-l KHz for the EEG and 8-10 KHz for the
round window response. The MGB and MRF channels were further high
pass filtered and all three channels were displayed on a Tektronix RM565
dual-beam oscilloscope. The EEG was recorded on an FM channel of
a Honeywell 7600 tape recorder and the high pass filtered MGB and
MRF activity was further amplified and recorded along with sync pulses
on three direct channels of the tape recorder. When the round window re-
sponse was monitored, it was also placed on a fourth direct channel. The
MGB and MRF channels were also led into amplitude discriminators
which produced a standard pulse each time the amplitude of the signal ex-
ceeded a present threshold.
340 IMIG, WEINBERGER, AND WESTENRERG
Clicks were the most frequently employed stimulus and were generated
from a 0.5 msec pulse produced by a Tektronix 161 pulse generator. A
Grason-Stadler 455C noise generator produced white noise and a \Vave-
tek 114 voltage-controlled oscillator provided pure tones. Either clicks,
white noise, or tones could be switched into both channels of a Grason-
Stadler electronic switch (model 829E) whose onset and offset were con-
trolled by Tektronix 161 and 162 modules. The rise-fall time for tone and
noise bursts was set for 5 msec. The output of each channel of the switch
was fed through Hewlett-Packard model 350D attenuators and into either
Grason-Stadler D30 or TDH49 earphones mounted at the end of each ear
bar. The stimulus was monitored with a Scott 260-B stereophonic amplifier
and two speakers mounted on each side of the equipment console. Stimula-
tion parameters (rate, frequency, duration, and intensity) were held con-
stant for each neuron which was studied. Stimulus intensities were approxi-
mately 85 db re .0002 pbar, measured by placing the earphone very close
to a Bruel and Kjaer 2205 impulse sound-level meter.
Following injection of Flaxedil the stainless-steel electrode was lowered
into the MRF. The glass-coated tungsten microelectrode was then lowered
by hand to a region just above the MGB. After this time, the electrode
was lowered in small steps by an electric drive mounted on top of the elec-
trode carrier and controlled by the experimenters outside the acoustic
room. Rhythmic acoustic stimulation was presented at a rate of approxi-
mately 1-2/set as the electrode was lowered. Unit waveform was moni-
tored carefully. Only neurons whose waveform was at least three times
larger than anything else in the record were studied. If the signal to noise
ratio deteriorated to the point that there was any uncertainty with respect
to what was being passed through the amplitude discriminator, data collec-
tion was terminated.
After a neuron was isolated, acoustic stimuli were delivered in blocks of
48-100 (“stimulation” blocks), alternating with equal intervals of silence
(“spontaneous” blocks). After collecting several blocks of data, a shock
train of S-10 shocks at approximately 2/set was delivered to the forepaw,
following a spontaneous block. Immediately after the shock, a stimulation
block was started and stimulation and spontaneous blocks alternated as be-
fore. The administration of shock was repeated if time permitted.
During the experiment, sync pulses and the standard output pulses from
the MGB channel amplitude discriminator were led to a LAB-S com-
puter. The computer timed the occurrence of the sync pulses and spike
pulses to the nearest 100 ysec and recorded the times on digital tape. This
information was subjected to further computer analysis after the experi-
ment. The data analyses are described in the results section.
At the termination of the experiment, marking lesions were made by pass-
&IEDI.iL GENICULATE NUCLEUS 311
ing 3-5 millicoulombs of current through the electrode tips and the animal
was killed with an overclose of Surital. The head was perfused with saline
solution followed by 10% formalin injected through the carotid arteries.
Serial coronal sections ( 50 :I.‘) were prepared on the freezing microtome
and every other section was mounted and stained alternately with cresyl
violet and \“\‘eil methods.
Results
Loclts of Cells. The classical division of the MGB into parvocellular and
magnocellular regions has been refined by the Golgi studies of Morest
(13) who suldivides the lateral parvocellular region into dorsal and ventral
components. It appears that only the ventral portion receives a strong
input from the brachium of the inferior colliculus and sends massive affer-
ents to the classical auditory cortex Nevertheless, auditory stimulation
(e.g., clicks 1 drive cells in both the dorsal and ventral regions. While it
would be of considerable interest to determine whether or not cells in the
two regions of the MGB differ in their functional characteristics, the pres-
ent study provides no such data. This deficiency results from the fact that
the two divisions are most easily identified in Golgi preparations and are
difficult, if not impossible, to differentiate in standard Nissl stains which
we employed. From a comparison of Morest’s figures (9) with our recon-
structions, there seems to be no question that the cells in this study lay
within both the dorsal and ventral regions of the MGB. In terms of stand-
ard stereotaxic coordinates they were between AP 3 and 5, and ML 10.5
and 12.0. However functional comparisons were not attempted becauseo f
the uncertainty of defining the regions in question in our material.
Ozw-a,11 Rate of Firing. The range of mean rates for all 20 cells was
0.5-lS/sec during both stimulation and silence. These figures were calcu-
lated over the entire period during which a cell was held, from 45 nun to
more than 2 hr. Such an estimate does not reveal the variability in rate
which might he exhibited by a single cell from time to time. Therefore,
a more detailed analysis was undertaken.
Measuvrmcnt of Spike Corrnts. The measurement of spike counts, i.e.,
the number of discharges per unit time, is one means of characterizing the
discharge of a single neuron. The time interval over which the spikes are
counted is delimited in this esperiment by two consecutive sync pulses
which are separated by 0.525-1.030 sec. When acoustic stimulation is pre-
sented, the sync pulses occur simultaneously with the stimuli; during the
collection of spontaneousa ctivity, sync pulses occur at the same rate as dur-
ing stimulation, but simply serve to delimit the intervals for spike count
computations.
Since the spike count is the unit of measurement which will be sub-
342 IJIIG, WEINBERGER, ASD \i’ESTEiXBERG
jetted to statistical analysis, its distribution is important for the selection
of appropriate statistical tests. The distribution of spike comets depends not
only on the statistical properties of the spike train from which it was de-
rived, but also on the time interval upon which the spike count is based.
For very short intervals, e.g., 1 msec, no more than one spike at most
could generally occur and therefore the distribution of spike counts would
be composed entirely of the categories 0 and 1. For longer intervals. a nor-
mal distribution might be approached.
Figure 1 shows some typical distributions of spikr counts. These distri-
butions were compiled from data which was col!ectecl during auditory
stimulation, but similar results obtain for spontnlV.:ous activity. The ah-
A
B
C
FIG. 1. Frequency distributions of spike counts for three cells. The abscissa dis-
plays the category of spike count (i.e., the number of discharges occurring per each
530 msec time interval) and the ordinate displays the frequency of occurrence of a
category. All spike count samples were collected during click stimulation. P refers
to samples taken prior to shock, S refers to samples taken after shock. A. Unit
71-23-4. Horizontal calibration = 69.2 ; vertical calibration L=I 25.4 ; n = 600 during P ;
IZ = 300 during S. B. Unit 71-26-3. Horizontal calibration = 6.4: vertical calibra-
tion = 102; II = 600 during P; n = 400 during S. C. Unit 71-23-3. Horizontal cali-
bration = 26.0; vertical calibration = 25.4 ; II = 600 during P ; ~1 = 600 during S.
343
MEDIAL GENICULATE NUCLEUS
scissa displays categories of spike counts (i.e., 0,1,2, . . . n where n is the
greatest number of spikes occurring in an interval) and the ordinate dis-
plays the frequency of occurrence of a category of spike count in the sam-
ple spike train. The spike count distribution in Fig. 1A was compiled from
unit 71-23-4. The distribution is clearly bimodal both prior to and follow-
ing shock. Figure 1B shows two distributions of spike counts both of
which were compiled from unit 71-26-3 prior to (P) and following (S)
paw shock. Like the preceding unit, the control distribution is bimodal;
however the postshock distribution from the same unit is unimodal. Figure
1C shows control anr’ postshock distributions from unit 71-23-3 and the
distributions are un&>del under both conditions. In addition to the occur-
rence of bimodal distributions, most of the distributions appeared to be
skewed to the left, a feature which is apparent in Fig. 1. It is clear that the
distributions of spike counts for these data are not normal ; therefore, non-
parametric statistics have been employed in all analyses of spike counts.
Stationurity of Rntc. The first question we sought to answer was
whether or not unit discharge rate changed significantly during the period
of time in which the unit was studied. In the sense used here, “stationar-
ity” refers to the absence of significant changes in rate over time for both
the stimulated and spontaneous conditions. The sequential distribution of
spike counts was examined for stationarity ; its display is termed the “se-
rial counts histogram” (SCH) .
An SCH is constructed by plotting the number of counts between con-
secutive clicks (or sync pulses) as a function of the serial order of the
stimulus. A stationary rate would appear as spike count values varying
about an unchanging mean value. The SCH of three units during both
spontaneous and stimulated conditions prior to and following shock are
presented in Fig. 2. Although spontaneous and stimulated treatment blocks
were alternated during the experiments, SCH are plotted separately for
consecutive spontaneous and stimulated conditions for ease of viewing the
cell’s discharge rate within a condition. Each SCH is divided into seg-
ments of equal length each of which represents a single data block (e.g.,
100 consecutive clicks).
Examination of Fig. 2 reveals considerable variability in the SCHs for
different neurons. Thus Fig. ZA depicts the SCHs of a cell which ap-
peared to be stationary in all conditions; although moment-to-moment var-
iability is seen, these appear random and no trends are evident. Nonsta-
tionarity is seen in Fig. 2B; there appears to be an increase in rate during
the preshock period of both spontaneous and evoked activity. Yet a third
pattern is seen for a third unit in Fig. 2C; a transient increase in rate oc-
curs following shock.
Instead of relying on the appearance of the SCH to subjectively evaluate
344 IMIG, WEINBERGER, AND WESTENBERG
the presence or absence of a change in discharge rate, the Kruskal-Wallis
(K-W j one way analysis of variance by ranks was used. Each data block,
containing 48-100 spike-count measurements, constituted one group in the
analysis; however in two units it was necessary to subdivide data blocks to
produce a sufficient number of groups for the analysis. If the K-W statis-
S
SPON
A
SPON
B
SPON
C
STIM
FIG. 2. Sequential spike count histograms for three cells. The abscissa displays
sequentially occurring intervals of time ; the number of discharges occurring during
each interval is displayed on the ordinate. P = spike count histogram assembled from
spike trains recorded prior to shock. S Z= spike count histogram recorded after paw
shock. SPON = no click stimulation presented. STIM = during click stimulation.
Divisions below SCH represent data blocks (see text). Horizontal calibration = 100
spike count values or 53 set (spike count interval = 530 msec). Vertical calibrations :
A. = 14.0 spikes; B. = 12.7 spikes; C. = 11.8 spikes.
MEDIAL GENICULATE NVCLEUS 345
tic was significant at the 0.05 level or less, the rate was classified as non-
stationary; otherwise the discharge rate was classified as stable or station-
ary.
In only four of the 20 units which were studied was the rate stationary
over the entire period during which the unit was recorded. Figure 2A il-
lustrates an example of one unit which does not appear to show a change
in rate during any period and the K-W test verifies the observation. But
for the majority (SO%) of the units, a significant change occurred at
some time during the analysis (e.g., unit 71-22-G in Fig. 2B).
Since the animal was given one or more paw shocks while the unit was
being recorded, it is conceivable that the nonstationarity was due entirely
to paw shock (i.e., a change in arousal). If the shock were the only varia-
ble which produced a significant change in the spike counts, then the first
control period, (i.e., prior to shock presentation) should be characterized
by stable discharge rates. But, of the 16 units which showed nonstationary
stimulated discharge rate over-all, seven (43% ) showed nonstationarity
during this control period.
Spontaneous activity sufficient for statistical analysis was collected in 14
units; three were stationary under all conditions. Of the 11 units which
showed significant change in spontaneous rate over-all, seven ( 63%)
showed significant change prior to the occurrence of paw shock. Thus. sta-
tistically significant changes occurred in the discharge rate of a significant
proportion of the units independent of any intentional change in the ani-
mal’s sensory environment or physiological state.
The Efect of Paw Sizocfi o+aD ischarge Rate. Following collection of
control data, shock was administered to the contralateral forepaw and data
collection was resumed. If the unit was held for a sufficient period. this
procedure was repeated one or more times. Since acoustic stimulation and
data collection were suspended only briefly during the 3-5 set required to
administer shock, any after effects of shock should be seen in the immedi-
ate postshock block which was recorded during the minute or so following
shock. Insofar as the purpose of shock was to produce a change in orga-
nismic state, it would be ideal to have an independent state measure. In
this regard, the electrocorticogram routinely led from the middle supra-
sylvian gyrus provided little information. Under the conditions of this ex-
periment the EEG was consistently desynchronized: slow waves or spin-
dles were never observed during the frequent periods of monitoring. Also,
units never exhibited bursting at approximately lO/sec which is typical of
thalamic cells during light sleep. This conclusion must be tempered by the
fact that quantitative analysis of the EEG was not attempted in this esper-
iment. Nevertheless, objective indices of state changes in these animals
were available. First, paw shock produced pronounced pupillary dilatation
346 IMIG, WEINBERGER, AND WESTENBERG
in nonatropinized animals. Second, it often produced prolonged (l-2 min)
increases in multiple-unit activity recorded from the mesencephalic reticu-
lar formation. We take these measures to indicate that the effect of paw
shock was generally an increase in arousal level superimposed upon the
waking or alert state.
A transient increase in rate often occurred following pawshock for both
spontaneous and stimulated conditions (Fig. 2C), however transient de-
creases in rate were never observed. Instead of relying upon visual inspec-
tion of the SCH to determine the effects of shock on spike rate, a statisti-
cal analysis was performed. The spike counts occurring during last
preshock and first postshock blocks for spontaneous and stimulated condi-
tions respectively, were compared by the Mann-Whitney U test. For the
stimulated condition, statistically significant changes in rate (p < .05)
were found for 11 out of 20 cells (55%). Nine of these showed an in-
crease in rate, one a decrease, and one both directions of change on succes-
sive shocks. The 20 cells received a total of 39 shocks. While 64% of these
tests (25 out of 39) revealed no significant rate change, tests in which the
rate did change were predominantly increases in ten out of 14 (71%).
Sufficient data with respect to the spontaneous condition were gathered for
14 cells. Rate changes were found for 8 out of 14 cells (57%), five show-
ing an increase, and three a decrease. These 14 cells received a total of 23
shocks. As for the stimulated condition, there was no change for most tests
(61%, 14 out of 23), but of the changes that did occur, the majority was
an increase in rate (67%, 6 out of 9).
In summary, paw shock effected the rate of discharge of slightly more
than half the cells. When a change did occur, it was quite likely to be an
increase in rate.
Transient increases in rate were sometimes observed to occur “sponta-
neously,” without the intervention of shock. For example unit 71-30-3
showed such an increase during block 2 of the preshock period (Fig. 3).
Similar findings are also evident for cell 71-30-2 (Fig. 6A) while high
rates during the preshock period were seen in blocks l-3 for unit 71-26-4
(Fig. 4). The possible importance of these spontaneous rate increases will
be discussed below in relation to the shape of the poststimulus histogram.
Patterns of Unit Discharge. The characteristic pattern of most cells con-
sisted of a short latency (< 20 msec) discharge following acoustic stimula-
tion, and in several cases a longer latency (90-l 10 msec) discharge. Later
peaks were evident in the post stimulus time histograms (PSTH) of only
a few cells. For one cell, the initial peak had a latency of 175 msec. This
particular cell was stimulated with a white noise burst whose offset oc-
curred at 160 msec so that this response might have been an “off” dis-
charge; this is conjectural because stimulus duration was not varied. Fol-