Synaptic interactions between primate precentral cortex neurons revealed by spike-triggered averaging of intracellular membrane potentials in vivo

Abstract

To document synaptic interactions between neurons in the precentral cortex of macaque monkeys, we recorded in vivo the intracellular (IC) membrane potentials of cortical neurons simultaneously with extracellular (EC) action potentials of neighboring cells. The synaptic potentials correlated with EC spikes were obtained by spike-triggered averages (STA) of the IC membrane potentials for 373 cell pairs recorded in anesthetized and awake behaving monkeys. Sixty-three STAs (17%) showed excitatory postsynaptic potentials (EPSPs), beginning after the trigger spike. Pure EPSPs had onset latencies of 0.9 +/- 0.7 msec (mean +/- SD) and amplitudes of 226 +/- 130 microV. Sixteen STAs (4%) showed postspike inhibitory postsynaptic potentials (IPSPs), with onset latencies of 0.4 +/- 0.4 msec and amplitudes of -274 +/- 188 microV. The most common waveform, observed in 82% of the STAs with features, was a broad depolarization straddling the trigger spikes, reflecting synchronized synaptic input to both IC and EC neurons. These average synchronous excitation potentials (ASEPs) began 14.3 +/- 6.6 msec before the trigger spike and had amplitudes of 1064 +/- 867 microV. Twenty-three STAs (6%) showed an average synchronous inhibitory potential (ASIP): a hyperpolarization beginning before the trigger spike and reflecting IPSPs produced by a group of local inhibitory cells synchronized with the trigger cell. ASIPs had an onset latency of -5.5 +/- 2.7 msec and amplitude of -589 +/- 502 microV. Combinations of synchronous and postspike potentials were also observed. Successive recordings provided examples of convergent and divergent connections between EC and IC cells. Neuron pairs with depolarizing postsynaptic potentials (PSPs) in the STA yielded peaks in the cross-correlograms of the IC and EC action potentials; the peak area was proportional to the amplitude of the PSP. These data suggest that a significantly larger proportion of cortical neurons interact through synchronous activity than through simple serial interactions; moreover, synchronous excitation affected more widely separated cell pairs than EPSPs and IPSPs, which were seen most often among the closest cells.

Fig. 1.

Schematic of cortical recording arrangement. Amplified signals obtained from IC electrode (IC) and EC electrode (EC) were recorded on tape and led to window discriminators (WD) and to a computer for STA. The only EC pulses accepted for STA were those occurring in the absence of IC pulses (like the two indicated by arrows). Recorded data were analyzed in a similar manner off-line. To facilitate generation of EC action potentials, Na-glutamate could be iontophoretically applied through a second barrel (Glu). (Note: The schematic drawings of the cells in this figure and Figure 4 should not be interpreted as implying particular cell types.)

Fig. 2.

Cortical sites of EC and IC recordings.A, Surface view of the penetration sites of EC and IC electrode pairs. Eight hemispheres of four monkeys are superimposed on a plane, in which penetration sites of the electrode pairs are marked as dots. Data from other monkeys were recorded from similar locations. B, Parasagittal section of a recording site (marked by an asterisk inA). The EC electrode track was indicated by blood cells and glia. C, Drawing of the recording sites, derived from B. Straight lines indicate the EC and IC electrode tracks; dots along tracks show sites of the recorded cells. Arrows indicate the pair whose STA is illustrated in Figure 4B.

Fig. 4.

Four basic types of synaptic potentials revealed by STAs. Two examples are illustrated for each type (top). Averages show triggering EC spikes (top traces) and averaged IC membrane potentials (bottom traces). The number of sweeps in each STA is shown inparentheses. Possible synaptic circuits mediating effects are diagrammed at the bottom. Pyramidal-shaped units represent IC cells; the rest, EC cells. Open boutons designate excitatory synapses; solid boutons, inhibitory synapses; shaded boutons, either excitatory or inhibitory synapses.

Fig. 3.

Examples of EC and IC recordings and STAs.A, Simultaneous recording of EC cell spikes and IC membrane potential. Two different EC units, with large and small action potentials, were spontaneously active under the anesthetized conditions. B, Schematic cross-section of cortex showing the IC and EC recording sites. C, STAs compiled from action potentials of the larger unit (L-u) showing an EPSP. Trigger spikes occurred at beginning of sweep for this figure and Figures 9 and 10. STAs compiled from the smaller action potentials (S-u) did not show any significant membrane potential deflections in the same numbers (n = 128) of sweeps. A control average of the same IC membrane potential during this recording period triggered from pulses generated regularly at 20 Hz did not produce significant deflections (Ctrl).

Fig. 5.

Amplitude distributions of synaptic potentials recorded with methylsulfate electrodes.

Fig. 6.

Onset latencies of synaptic potentials relative to onset of trigger spikes.

Fig. 7.

Rise times (0–100%) of synaptic potentials plotted against amplitudes.

Fig. 10.

Divergent output effects from two EC cells to multiple IC cells. Activities of two EC units (action potentials atbottom left) were recorded from the same location (asterisk, top left). The STA from the larger EC unit showed an IPSP in the fourth IC neuron (middle). Hyperpolarizing current reversed the polarity of this IPSP. a, Control; b, reversed potential with 1.0 nA; c, potential with 2.0 nA. STAs from the smaller EC unit (right column) showed EPSPs in the third and fourth IC neurons. With hyperpolarizing current, this potential increased in amplitude. a, Control;b, 1.0 nA; c, 2.0 nA.

Fig. 8.

Combinations of synchronous and serial synaptic potentials. A, Two examples of an EPSP superimposed on ASEP. The post-trigger rise of the potential at arrowindicates onset of EPSP. B, Two examples of IPSP (onset at arrow) superimposed on ASIP. C, Two examples of IPs superimposed on ASEP. These records were obtained with two simultaneously recorded EC cells converging on the IC unit. The cross-correlogram of the EC cells (bottom) confirms their synchronous activation.

Fig. 9.

Convergent inputs from multiple ECs to the same IC cell. A, Schematic drawing of the EC and IC electrode tracks within the cortex and their recording locations (left) and the averaged potentials obtained by STA from each EC point. B, When the horizontal distance between the electrodes was >1 mm, serial synaptic interactions were rarely observed. C, When the distance was very close (0.2 mm), many EC cells showed EPSPs with various time courses.

Fig. 11.

Amplitudes of synaptic potentials plotted against separation of EC and IC electrode tips.

Fig. 12.

Relationship between synaptic potentials and cross-correlograms. Examples (top) illustrate correlograms between EC and IC cells associated with EPSP (left) and ASEP (right). STAs were compiled in absence of IC spikes. Graph plots correlogram peak area (number of above-baseline counts in correlogram peak per trigger spike) against amplitude of synaptic potential in the STA. Linear regression line has slope of 0.3 spikes/mV.