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Dynamics of Depolarization and Hyperpolarization in the Frontal Cortex and Saccade Goal

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Science  01 Feb 2002:
Vol. 295, Issue 5556, pp. 862-865
DOI: 10.1126/science.1066641

Abstract

The frontal eye field and neighboring area 8Ar of the primate cortex are involved in programming and execution of saccades. Electrical microstimulation in these regions elicits short-latency contralateral saccades. To determine how spatiotemporal dynamics of microstimulation-evoked activity are converted into saccade plans, we used a combination of real-time optical imaging and microstimulation in behaving monkeys. Short stimulation trains evoked a rapid and widespread wave of depolarization followed by unexpected large and prolonged hyperpolarization. During this hyperpolarization saccades are almost exclusively ipsilateral, suggesting an important role for hyperpolarization in determining saccade goal.

Electrical stimulation has been used extensively in brain research since its introduction more than 100 years ago (1). Despite the popularity of this technique and its potential clinical applications (2, 3), many questions remain regarding the spatial profile and the temporal dynamics of the neural response that is evoked by intracortical microstimulation (4, 5). One of the first cortical regions that was systematically explored using microstimulation is the frontal eye field (FEF) (6–8), a cortical area situated in the anterior bank of the arcuate sulcus and known to be involved in planning and execution of saccadic eye movements (9–11). The FEF and neighboring area 8Ar (12) contain a representation of saccades to the contralateral hemifield. Microstimulation in these regions evokes short-latency conjugate saccades that depend on the stimulation site and are very similar to voluntary saccades (6–8).

We sought to measure and characterize directly the neural response that is elicited by microstimulation in the frontal cortex of the behaving monkey and to determine the relation of this response to the saccadic eye movements that are evoked by stimulation. We thus combined standard electrophysiological techniques in awake behaving monkeys with optical imaging using voltage-sensitive dyes (13). This combination provides a powerful paradigm for measuring the electrical activity of populations of neurons in the brain of behaving monkeys at excellent spatial and temporal resolutions (14).The dye signal measures the sum of the membrane-potential changes in all the neuronal elements in the imaged area, emphasizing subthreshold synaptic potentials in neuronal arborizations originating from neurons in all cortical layers whose dendrites reach the superficial cortical layers (15).

Combined microstimulation (16) and optical imaging were applied to the frontal cortex in four hemispheres of two monkeys at a total of 39 recording sites (17) while the monkey performed a simple fixation task (18). To image the FEF and 8Ar, we opened a cranial window over the arcuate sulcus, then removed the dura and replaced it with a transparent artificial dura (19). The imaged/stimulated area included primarily a narrow strip of cortex, lying 1 to 3 mm anterior to the arcuate sulcus, from which short-latency contralateral saccades could be evoked (16, 20). Figure 1A shows the frontal cortex in the left hemisphere of one monkey 3 weeks after surgery; the arcuate sulcus (Ar) and the principal sulcus (Pr) can be seen through the transparent dura. We stained the cortex with blue voltage-sensitive dyes and imaged the resulting activation pattern at up to 400 frames per second (13).

Figure 1

Spatiotemporal dynamics of microstimulation-evoked activity. (A) The FEF in the left hemisphere as seen through the cranial window and the transparent artificial dura. The dorsomedial arm of the arcuate sulcus (Ar) can be seen, as well as the principal sulcus (Pr). The area imaged in the experiment shown in (B) is marked by a white rectangle. (B) Sequence of images of the cortex during microstimulation. The time interval between frames is 9.6 ms. The imaged area is 6.8 mm by 6.8 mm. Stimulation is applied through frames 2 to 4 and is indicated by the black horizontal line. The site of penetration of the stimulating electrode is marked by an X. Stimulation at this site evoked a 30° saccade to the right. The optical signals were low pass–filtered (σ = 250 μm) for display purposes only. The activation pattern is averaged across 30 repetitions.

The activation pattern evoked by microstimulation at one stimulation site is shown in Fig. 1B. A train of stimulation pulses (50 μΑ, 500 Hz) was applied for 24 ms at the site marked by X. Shortly after stimulation onset there was a large and widespread increase in fluorescence, which corresponds to an overall depolarization. This depolarization spread over a large cortical area in a rather uniform manner, consistent with the spread of activity measured in the visual cortex (21, 22). Following this brief depolarization, the response dropped sharply, and most of the imaged area became dark. This large and rapid drop in the optical signal corresponds to an overall hyperpolarization of the average membrane potential of the neuronal population to a level well below baseline (23).

The mean time course of the optical signal for four different stimulation conditions was measured in a small area 0.4 mm by 0.4 mm near the site of the stimulating electrode (Fig. 2). A short stimulation train of 24 ms (green curve) elicited a rapid depolarization that was followed by a large and prolonged hyperpolarization. The hyperpolarization returned to baseline about 230 ms after stimulation onset and showed a small depolarization overshoot. When stimulation was applied for a longer period of 80 ms (dark blue curve), the response still peaked at about 25 ms and then dropped to a lower depolarization level despite the continuation of the stimulation. When the train of stimulation was divided into four short trains with short gaps in between (red and cyan curves), the response went through four cycles of depolarization and hyperpolarization. This large hyperpolarization was typical for most stimulation sites in the frontal cortex. In contrast, in the primary visual cortex of the awake monkey, the same stimulation parameters do not evoke similar hyperpolarization (22).

Figure 2

Time course of the response to microstimulation. The curves show the mean ± SEM of the response to four stimulation conditions in a 0.4 mm by 0.4 mm area of cortex near the site of stimulation, measured from the same site as in Fig. 1. The number of repetitions in each condition is between 27 and 30. The horizontal lines at the bottom indicate the stimulation time for each stimulation condition.

The time course of the optical signal shows that each depolarization response in the frontal cortex is followed by a similar, and sometimes even larger, hyperpolarization response. How does this interplay between depolarization and hyperpolarization affect the saccades that are elicited by microstimulation? The horizontal eye velocity traces from three trials in which eye movements were evoked by stimulation at a site in the right frontal cortex that represents a 20° saccade to the left are shown in Fig. 3A. At this site and with these stimulation parameters there was a large variability in saccade initiation time, and in these three trials saccades were initiated 50, 80, and 100 ms after stimulation onset. Although in all three trials saccades were evoked under identical stimulation conditions, these saccades differ markedly. The early contralateral saccade (red curve) displayed the maximal amplitude and peak velocity characteristic for this site and was followed 130 ms later by a large ipsilateral saccade. The saccade that started around stimulation offset (green curve) displayed a smaller amplitude and reversed to the ipsilateral direction at midflight. The saccade that started about 20 ms after stimulation offset (cyan curve) was short and ipsiversive. All of these eye movements were evoked by the stimulation; on randomly interleaved trials without stimulation, the monkeys made no saccades during this interval.

Figure 3

Examples of the correlation between the properties of stimulation-evoked saccades and the response elicited by microstimulation. (A and C) Horizontal eye velocity traces for three trials with 80-ms stimulation (A) or four trains of 24-ms stimulation separated by an interval of 44 ms without stimulation (C). The horizontal lines at the bottom of each panel indicate stimulation times. (B and D) Comparison of the mean ± SEM of the time course of the optical signal (green curves) measured in a 0.4 mm by 0.4 mm area of cortex near the site of stimulation, and individual traces of horizontal eye velocity (blue curves) for long stimulation (B) or four short trains (D). Red curves indicate the mean ± SEM of horizontal eye velocity on nonstimulated trials (n = 52).

The changes in saccade properties over time closely parallel the time course of the optical signal measured near the stimulation site. The mean time course of the optical signal (green curve) and the horizontal eye velocity traces (blue curves) for all the trials at this site and at these stimulation parameters (n = 25 trials) are shown in Fig. 3B. Saccades that were initiated during strong depolarization were contralateral and displayed high peak velocity. Saccade peak velocity decreased during the drop in depolarization and then reversed to the ipsilateral direction. Saccades that were initiated during strong hyperpolarization were almost always large return saccades in the ipsilateral direction.

The correlation between the measured response and the properties of the evoked saccades holds also for other stimulation conditions.Figure 3C shows eye movements that were evoked during three trials of stimulation with four short trains at the same site as in Fig. 3, A and B. In these trials, the first short train failed to evoke a short-latency saccade. The first eye movement evoked in those trials was a short ipsiversive saccade rather than the large contraversive saccade typical for this site. In two trials (red and green curves), ipsiversive saccades were initiated about 35 ms after the offset of the second stimulation train, during the initial phase of hyperpolarization. During this ipsiversive saccade the third train was applied and evoked a large contraversive saccade. It was followed by another small ipsiversive saccade that coincided with another period of hyperpolarization. In the third trial (cyan curve), an ipsiversive saccade was initiated about 40 ms after the offset of the third train and showed the same sequence of ipsi-contra-ipsi movements.

Figure 3D compares the horizontal eye velocity traces (blue curves) and the mean optical response (green curve) for all trials with the same stimulation condition as in Fig. 3C (n = 26). The direction and peak horizontal eye velocities of the evoked saccades are highly correlated with the optical signal measured at the time of saccade initiation. The first two cycles of depolarization evoked almost no saccades. Saccades started preferentially during the hyperpolarization that followed the second or the third train and were almost always ipsiversive. During the periods of depolarization following the third and fourth trains the eyes moved toward the contralateral direction; during the hyperpolarization intervals following the second, third, and fourth trains the eyes moved toward the ipsilateral direction.

Following microstimulation, the level of depolarization or hyperpolarization in FEF and 8Ar at the time of saccade initiation proves to be an excellent predictor of the direction, size, and peak velocity of the evoked saccades across our entire data set. The results obtained from 39 stimulation sites and 1044 stimulation-evoked saccades are summarized in Fig. 4(24). Each panel shows a scatter plot of normalized saccade peak horizontal velocity against saccade initiation time. For this analysis we combined all saccades that occurred shortly after stimulation, irrespective of whether these were the first (red) or later saccades within a given trial (cyan). Under the two stimulation protocols there is a strong correlation between the measured optical signal and the properties of the evoked saccades. Saccades that were initiated during depolarization were almost exclusively contraversive. Saccades that were initiated during a period of hyperpolarization were almost exclusively ipsiversive. Saccades that were initiated when the absolute level of the response was high tended to have a high peak velocity and a large amplitude; saccades that started when the response was low tended to have a low peak velocity and a small amplitude (25). The correlation between the mean optical signal (green curve) and peak horizontal eye velocity of individual saccades (red and cyan x's) is highly significant for both stimulation conditions [r = 0.93 for continuous stimulation andr = 0.82 for four short trains; P < 10−6 for both conditions (26)].

Figure 4

Correlation between saccade peak velocity and the electrical response elicited by stimulation. (A) Long stimulation train. (B) Four short trains. The green curve shows the mean ± SEM of the normalized time course of the optical signal measured at all sites in which a reliable optical signal was obtained (17). The scatter plot of x's indicates the normalized peak horizontal eye velocity versus initiation time for all saccades obtained during this interval in all stimulation sites (24). Red x's indicate the first saccade evoked on a given trial, and cyan x's indicate later saccades. The blue line depicts the running average of the normalized peak horizontal eye velocities (in a bin of 10 ms). All of these saccades were taken from a period in which no saccades occurred during control trials without stimulation.

Our findings raise two questions regarding the hyperpolarization. First, what is the role of hyperpolarization? Hyperpolarization could mediate saccade target selection through competitive interactions between sites that represent alternative saccades (27). It could also serve as a reset mechanism necessary for normal saccade sequencing (9). Once a saccade of the desired amplitude and direction had been executed, activity that encodes this saccade should rapidly terminate so that another similar saccade would not be initiated erroneously. Indeed, many FEF neurons with presaccadic activity are actively suppressed following a saccade into their response field (28, 29), suggesting that hyperpolarization may occur also after voluntary saccades. Hyperpolarization could also facilitate a rapid return saccade that would bring the eyes back to the previous eye position after desired and undesired saccades. Such rapid “glances” also occur during natural visual scanning behavior. A second question concerns the source of the hyperpolarization. In many experiments we found that hyperpolarization and depolarization started from different locations and displayed different spatial profiles (e.g., Fig. 1B). These results suggest that part of the hyperpolarization is caused by a stimulation-evoked recurrent inhibitory signal rather than by the well-known after-hyperpolarization. The finding that during hyperpolarization saccades are preferentially ipsilateral suggests that some of the delayed inhibitory signals could be mediated by “push-pull” inhibitory interactions between sites that encode opposite saccades in the two hemispheres (30). An exciting possibility, therefore, is that when the stimulation site is hyperpolarized, the site that encodes the opposite saccade in the other hemisphere is depolarized, contributing to the ipsilateral saccade.

Finally, our results demonstrate that neural activity with complex spatiotemporal dynamics can be elicited by microstimulation; these dynamics depend on the stimulated area and can have important behavioral correlates. These findings emphasize the importance of further characterization of microstimulation-evoked activity for the interpretation of the behavioral effects of microstimulation. Our finding that during hyperpolarization saccades are almost exclusively ipsilateral imposes new constraints on models of the neural mechanisms that compute saccade goal. Specifically, our results suggest that suppression of neural activity at a certain location in the saccade motor map in the frontal cortex is interpreted by downstream saccade circuitry as a signal in favor of an ipsilateral saccade.

  • * To whom correspondence should be addressed. E-mail: eyal.seidemann{at}weizmann.ac.il

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