1Department of Integrative Physiology,
Aizawa, Hiroshi,
Yasushi Kobayashi,
Masaru Yamamoto, and
Tadashi Isa.
Injection of Nicotine Into the Superior Colliculus Facilitates
Occurrence of Express Saccades in Monkeys.
J. Neurophysiol. 82: 1642-1646, 1999.
To clarify the role
of cholinergic inputs to the intermediate layer of the superior
colliculus (SC), we examined the effect of microinjection of nicotine
into the SC on visually guided saccades in macaque monkeys. After
injection of 0.4-2 µl of 1-100 mM nicotine into the SC, frequency
of extremely short latency saccades (express saccades; reaction
time = 70-120 ms) dramatically increased, for the saccades the
direction and amplitude of which were represented at the location of
the injection site on the collicular map. However, no marked change was
observed for the relationship between the peak velocities and the
amplitudes of saccades. These results suggested that activation of
nicotinic acetylcholine receptors in the SC can facilitate initiation
but causes no major change in dynamics of visually guided saccades.
The mammalian superior colliculus (SC) is a
central structure controlling visually triggered behaviors such as
saccadic eye movements and orienting behaviors etc. (for review, see
Sparks 1986 It is well known that the SGI receives dense acetylcholinergic
innervation in various species of mammals such as rats, guinea pigs,
cats, ferrets, and primates (Beninato and Spencer 1986 Recently in slice preparations of the rat SC, we found that application
of an acetylcholinergic agonist nicotine induced inward currents and
depolarization in the SGI neurons by direct activation of nicotinic
acetylcholine receptors (nAChRs) (Isa et al. 1998b Behavioral studies on the saccadic reaction times (SRTs) have shown
that distribution of SRTs shows at least two distinct peaks: regular
saccades with SRTs of 150-250 ms and express saccades with SRTs ~100
ms (Fischer and Boch 1983 Two Japanese monkeys (Macaca fuscata) were fitted
with an acrylic skullcap and scleral search coils (Fuchs and
Robinson 1966 Effects of nicotine injection (0.4-2 µl, 1-100 mM) into the SC
on SRTs were examined in a total of 13 experiments in two monkeys (experiments N1-N13, see Table 1). As
control, injection of vehicle (saline) was tested in four experiments
(experiment S1-S4 in Table 1). Figure 1,
A-D, shows the result of the experiment N10, in which 2 µl of 10 mM nicotine solution was injected. As shown in Fig. 1,
A and B (NO GAP paradigm), the SRTs
were distributed primarily between 120 and 200 ms (mean ± SD;
163.8 ± 11.4 ms, n = 22) before the nicotine
injection (Fig. 1B,
ABSTRACT
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ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
; Wurtz and Albano 1980
). The
superficial layer of the SC (the stratum griseum superficiale; SGS)
receives visual input from the retina (Altman 1962
;
Lashley 1934
) and the primary visual cortex
(Harting et al. 1992
). The intermediate layer (the
stratum griseum intermediale; SGI) and the deep layer send descending motor command to the brain stem reticular formation and the spinal cord
(Huerta and Harting 1982
). As to the connection between
the SGS and SGI, recent studies by Lee et al. (1997)
and
our laboratory (Isa et al. 1998a
) showed the
electrophysiological evidences for existence of an excitatory
connection from the SGS to the SGI in the SC slices obtained from the
tree shrew (Lee et al. 1997
) and rat (Isa et al.
1998a
).
; Graybiel 1978
; Hall et al. 1989
;
Henderson and Sherriff 1991
; Jeon et al.
1993
; Ma et al. 1991
; Schnurr et al.
1992
). These acetylcholinergic fibers originate from the
pedunculopontine and laterodorsal tegmental nuclei (Beninato and
Spencer 1986
; Hall et al. 1989
). These
observations may suggest that the acetylcholinergic input to the SGI
modulates execution of visually triggered behaviors. Despite the
abundance of anatomic data, however, the functional implication of the
acetylcholinergic input to the SC has not been understood.
). We
have clarified further that activation of nAChRs in the SGI neurons
enhances signal transmission in the pathway from the SGS to the SGI
neurons. If this mechanism holds true also in primates, the consequence
of the facilitation may be direct activation of saccade-related burst
neurons in the SGI via the SGS-SGI pathway, behaviorally allowing the
immediate release of a reflex-like saccade toward a target.
; Fischer and Ramsperger 1984
). In the present study, we examined the effects of
microinjection of nicotine into the SGI in monkeys, especially the
effects on the SRTs, and found that activation of nAChRs in the SC
markedly reduced the SRTs and increased the frequency of express
saccades without changing their dynamics. The present results may
explain some of the neuronal mechanisms related to execution of express saccades and how nAChRs are involved in control of attentive behaviors.
METHODS
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ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
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) asceptically under halothane anesthesia. The
surgical procedures have been described in a preceding paper
(Aizawa and Wurtz 1998
). Animals were trained to perform
a visually guided saccade paradigm sitting in a primate chair with
their heads fixed. Visual stimuli were given as small light spots
(0.8 × 0.8° square) back-projected on a tangential screen at a
distance of 28 cm from the eye. Three different tasks were shuffled
randomly; NO GAP paradigm (the saccade target appeared at
peripheral visual field after central fixation), GAP
paradigm (the target was lit after 170 ms gap after the fixation offset), and catch trials (after the gap, the target was emitted at the
fixation point again). GAP and NO GAP paradigms
in opposite directions were presented with equal probability (22%,
respectively). Catch trials consisted 12% of the whole session. In all
the paradigms, the duration of central fixation was varied randomly
between 300 and 1,000 ms. Behavioral paradigms, visual displays, and
collection and storage of eye-movement data were under the control of a
real-time data-acquisition system (REX) (Hays et al.
1982
). Location of the tip of the microsyringe was identified
by the vector and threshold of saccadic eye movements evoked by
electrical stimulation (<20 µA, 500 Hz, 50 biphasic pulses of 200 µs width) via the electrode attached to the microsyringe (Crist,
MRM-S02) (see Aizawa and Wurtz 1998
). For the injection,
the areas in the SC that represent 5-20° saccades were selected. The
locations of the visual targets were determined to induce the saccades
with the same vector as those evoked at the injection site
("contraversive saccades"), and those of the same amplitude but on
the opposite side of the horizontal and vertical meridians
("ipsiversive saccades"). Contraversive and ipsiversive saccades
were shuffled randomly. The timing of saccade onset was determined at
the time when the eye velocity exceeded 30°/s. Saccades with reaction
time <70 ms were regarded as "anticipatory saccades," which were
not rewarded and discarded from data (see RESULTS).
Saccades with reaction time >350 ms were also not rewarded and
discarded from data. Frequency of anticipatory saccades did not exceed
3% of the trials in the present experiments. Either solution of
nicotine (1-100 mM in saline) or vehicle (saline) was injected at a
rate of 0.2 µl/min until the total volume reached 0.4-2 µl, and
reaction times, amplitude, and velocities of saccades were analyzed.
Experiments were performed in accordance with the NIH Guidelines for
the Care and Use of Laboratory Animals.
RESULTS
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ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
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). After nicotine injection, the SRTs
were shortened (146.8 ± 23.8 ms, n = 107 observed
during 30 min after injection) with a statistically significant
difference from those before injection (P < 0.05, Student's t-test). As clearly illustrated in Fig.
1A, the SRTs stepped down from the range of 150-200 ms to
extremely short range (<120 ms). The distribution of SRTs in the
present experiments thus were divided into two different subgroups at
120 ms (Fig. 1B) as in the preceding study (Dorris et
al. 1997
). Saccades with SRTs shorter than 70 ms were regarded
as anticipatory saccades and discarded from data. Therefore we
classified the saccades with SRTs between 70 and 120 ms as "express
saccades" and those with SRTs longer than 120 ms as "regular saccades" (Fischer and Boch 1983
). The increase in
frequency of express saccades continued for longer duration in the
GAP paradigm (Fig. 1, C and D) as
compared with the NO GAP paradigm (Fig. 1, A and
B). The average SRT of express saccades in the
GAP paradigm (100.1 ± 6.5 ms, n = 51)
was slightly shorter than that in the NO GAP paradigm
(104.8 ± 6.9 ms, n = 22). The difference was
statistically significant (P < 0.05, Student's
t-test) but minor. Thus the major change caused by
introducing GAP was the prolonged duration of the nicotine
effect.
Table 1.
Average SRTs for all the experiments
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Fig. 1.
A-D: effects of nicotine injection (10 mM, 2 µl,
experiment N10) into the monkey superior colliculus (SC) on saccadic
reaction time (SRT) during NO GAP and GAP (170 ms) visually guided saccade paradigms. Amplitude of the saccades was
7° in horizontal and 7° in vertical directions. Horizontal bars in
A and C indicate period of nicotine
injection (10 min). A and C: SRTs are
plotted against time. Open circles, trials before injection; asterisks,
trials during and after nicotine injection. A and
C, NO GAP and GAP paradigms,
respectively. B and C: histograms of the
normalized distribution of SRTs before (lower open columns) and after
application of nicotine (upper closed columns, 30-min recording) in
NO GAP (B) and GAP
(D) paradigms. E-H: effect of saline
injection (experiment S4). Amplitude of saccades was 10 deg horizontal.
Same arrangement as A-D.
As shown in Fig. 2, the average SRTs
(A and C) decreased [plots fell below the
equality line (slope = 1)], and the percentage of express
saccades (E and G) increased (the plots fell
above the equality line) for contraversive saccades both in NO
GAP (A and E) and GAP
(C and G) paradigms in a majority of nicotine
injections (). Statistically significant reduction in average SRTs
(P < 0.05, Student's t-test) was observed
in 12 of 13 experiments for GAP paradigm and 9 of 12 experiments in NO GAP paradigm as indicated by asterisks in
Table 1. The occurrence probability of express saccades showed
statistically significant increase after nicotine injection both in
NO GAP and GAP paradigms for the population data (n = 13, P < 0.05, Student's
t-test). As to the ipsiversive saccades, on the other hand,
no statistical change was observed in a majority of the cases.
Statistically significant change (P < 0.05, Student's
t-test) was observed in average SRTs in experiments N2, N8,
N9 (NO GAP) and N11, N12 (GAP; Table 1).
However, no consistent tendency was observed as to whether there was
significant increase or decrease in average SRTs and in occurrence
probability of express saccades (see also Fig. 2, B, D, F,
and H). Thus the effect of nicotine was spatially selective.
Injection of saline caused no significant effect in a majority of the
cases as shown in Fig. 1, E-H (see the population data in
Fig. 2) except for the experiment S3 in which the SRTs decreased for
contraversive saccades in both GAP and NO GAP
paradigms and experiment S4 in which the average SRTs increased for
ipsiversive saccades in GAP paradigm (P < 0.05, Student's t-test; Table 1).
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In addition, the effect of nicotine was dose dependent. When the effects of two different doses of nicotine into the same location of the SC were compared [0.4 µl of 50 mM (experiment N11) and 0.4 µl of 100 mM (N13)], the effect of the larger dose was longer in duration; however, there was no significant difference in the SRTs of the express saccades (104.6 ± 6.9 ms in N11 and 101.6 ± 11.0 ms in N13, P = 0.12, Student's t-test).
Figure 3 shows the relationship between the amplitude and peak velocity of eye movements during express saccades before (A and B) and after nicotine injection (C and D) in NO GAP and GAP paradigms (data from 7 experiments in 1 monkey). Linear regression lines between the amplitude and peak velocity virtually superimposed before and after the nicotine injection for both NO GAP and GAP paradigms (Fig. 3, B and D). The difference was not statistically significant for NO GAP paradigm (P = 0.10, Student's t-test) but significant for GAP paradigm (P = 0.04, Student's t-test). In the latter case, however, the difference was apparently minor (change in slope of the regression line was 3%, Fig. 3D). Thus nicotine injection increased the frequency of express saccades but caused no major change in their dynamics.
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DISCUSSION |
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Schiller et al. (1987) showed that express saccades
never occurred after ablation of the SC and concluded that the SC is an essential structure for execution of express saccades. Recent electrophysiological studies showed that express saccades are generated
when the visual response of saccade-related burst neurons in the SGI
triggers the motor burst (Dorris et al. 1997
;
Edelman and Keller 1996
). Dorris et al.
(1997)
further showed that gradual increase in activity in a
population of SGI neurons (buildup neurons) before the target
appearance was correlated significantly with the occurrence of express
saccades. Thus advanced increase in excitability level of the burst
neurons may lead to lowering the threshold for trigerring the motor
burst activity by a visual response, which leads to execution of
express saccades. In contrast, when the advanced increase in activity
is not sufficient, the visual response cannot trigger the motor burst;
this results in regular saccades. Recently we showed that activation of
nAChRs can induce inward currents and depolarization in a majority of the SGI neurons in slice preparations of the rat SC (Isa et al. 1998b
). Furthermore, depolarization of the SGI neurons induced by nicotine lowered the threshold for bursting response of the SGI
neurons to electrical stimulation of the SGS. Thus activation of nAChRs
could facilitate signal transmission in the direct visuomotor pathway
in the SC, the existence of which we proved in the rat SC slices
(Isa et al. 1998a
). The origin of the visual response in
the SGI neurons of monkeys is not yet clear; however, these studies
have suggested that express saccades can be generated by gating of
direct signal transmission from the SGS to the SGI. In the present
study, we observed that both introduction of GAP and
increase in nicotine dose did not cause further reduction in the SRTs
of express saccades. A slight increase in eye velocities was observed
after nicotine injection for the GAP paradigm (Fig. 3D); however, the increase was much smaller than the effect
caused by injection of GABAA antagonist bicuculline into
the SC, which also increases frequency of express saccades
(Hikosaka and Wurtz 1985
). Thus no major change was
observed in the dynamics of express saccades after nicotine injection.
These results in part support the Fischer's three-loop model in which
he proposed that that the loop with the shortest transmission time was
used for express saccades (Fischer 1987
).
Another interesting observation in the present study is that effect of
nicotine injection was more prominent in the GAP paradigm than in the NO GAP paradigm even though they were randomly
shuffled in the same block of trials. Thus the threshold for induction of express saccades by nicotine appeared to be lower in the
GAP paradigm. This "GAP effect" may be
partly caused by decrease in activity of fixation cells in the rostral
pole of the SC, which can suppress initiation of saccades, during the
gap period (Dorris and Munoz 1995). If so, the
cholinergic system can facilitate induction of express saccades
independently of release from inhibition by the fixation cells.
Besides the decrease in activity of fixation cells, advanced increase
in activity of buildup cells (buildup activity) is correlated with
occurrence of express saccades (Dorris et al. 1997).
However, the origin of the buildup activity during the GAP
period is still unknown. Furthermore it has been shown that frequency
of express saccades increases with daily practice (Fischer and
Ramsperger 1986
). A question worth further investigation is
whether the cholinergic input to the SGI originated from the
pedunculopontine and laterodorsal tegmental nuclei is involved in the
factors that facilitate occurrence of express saccades, such as buildup
activity of the SGI neurons and daily practice.
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ACKNOWLEDGMENTS |
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We thank Y. Takeshima, C. Suzuki, J. Yamamoto, and C. Kamada for technical assistance.
This study was supported by grants from the Ministry of Education, Science, Sports, and Culture of Japan to T. Isa (08458266, 08279207, and 09268238), CREST of the Japan Science and Technology Corporation, the Daiko Foundation, Naito Memorial Foundation, Uehara Memorial Foundation, and Mitsubishi Foundation.
Present address of H. Aizawa: Dept. of Physiology, School of Medicine, Hirosaki University, Zaifu-Cho 5, Hirosaki 036-8562, Japan.
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FOOTNOTES |
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Address for reprint requests: T. Isa, Dept. of Integrative Physiology, National Institute for Physiological Sciences, Myodaiji, Okazaki 444-8585, Japan.E-mail: tisa{at}nips.ac.jp
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Received 13 January 1999; accepted in final form 3 June 1999.
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