Formation of long-term potentiation in superior colliculus slices from the guinea pig

Neuroscience Letters, 96 (1989) 108 113 Elsevier Scientific Publishers Ireland Ltd.

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NSL 05806

Formation of long-term potentiation in superior colliculus slices from the guinea pig Yasuhiro Okada and Takaaki Miyamoto Department of Physiology, School of Medicine, Kobe University, Kobe (Japan) (Received 18 July 1988; Revised version received 12 September 1988; Accepted 12 September 1988)

Key words: Superior colliculus slice; Tetanic stimulation; Long-term potentiation; N-Methyl-D-aspartic acid receptor. Superior colliculus slices of sagittal section were prepared from the guinea pig. Postsynaptic potential (PSP) was evoked in the superficial grey layer (SGL) after the electrical stimulation to optic layer. Tetanic stimulation to the optic layer elicited long-term potentiation (LTP) in the PSP of the SGL. Tetanic stimulation of 20 s in duration and 50 Hz in frequency was most effective for the formation of LTP. The LTP formation was masked during application of 2-amino-5-phosphonovalerate (APV), a specific antagonist for N-methyl-D-aspartate (NMDA) receptor, but LTP was observed when APV was removed from the perfusion medium.

Since the discovery of long-term potentiation (LTP) by Bliss and Lomo in 1973 [2], research findings on the formation of LTP in the mammalian brain have mainly come from the hippocampus, and the neocortex areas [14]. LTP formation in the superior colliculus (SC) has not been reported in the literature, although LTP has been recorded in the goldfish optic tectum [9]. In this paper we report the first demonstration of LTP formation in SC slices from the mammalian brain. SC was chosen because its electrophysiology, biochemistry and pharmacology have been widely studied either in vivo using whole-brain preparations or in vitro in slice preparation [1, 6, 8, 11, 12]. It has been shown [7, 10, 14, 15] that LTP formation can be mediated by the N-methyl-D-aspartate (NMDA) receptor, a subtype of the glutamate receptors. In hippocampus, it has been reported [14] that the antagonists of glutamate receptors such as L-glutamate diethylester (GDEE), ~-D-glutamylglycine (7DGG) and 2-amino-5-phosphonovalerate (APV), inhibited the formation of LTP. In this experiment, we also report that APV, a specific N M D A receptor antagonist, blocked the appearance of LTP but after washing out of the agent, LTP was observed. Guinea pigs weighing between 200 and 300 g were decapitated, and the brains Correspondence: Y. Okada, Department of Physiology, School of Medicine, Kobe University, Kusunokicho, Chuo-ku, Kobe 650, Japan.

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quickly removed. Tissue blocks of SC were rapidly dissected out and cut sagittaUy (see Fig. 1A) into slices of between 400 and 600 pm thickness with a razor blade. Before starting the experiments, the slices were preincubated for a minimum of 20 min in the standard medium (in mM: glucose 10, NaCI 125, KC1 3, KH2PO4 1.24,

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Fig. 1. A schematic drawing of the sagittal cutting of the superior colliculus, the placement of the stimulating and recording electrodes and an example of LTP formation in a single slice of the SC when the tetanic stimulation (at 50 Hz for 20 s) was applied. A: a block of the brainstem containing the superior and inferior colliculus. The superior colliculus was cut sagittaUy into half at the centre. Five to 6 slices were obtained from each SC. It is important that cutting must be done slightly oblique using the fibre input of the optic nerve as a guide. Cross-section cutting of the SC does not result in good recording of PSP. B: the arrangement of the recording and stimulating electrodes. C~: two kinds of negative potentials in the control response. Note the earlier deflection (0 in the declining phase of the large potential(s). C2: 10 min after removal of Ca 2+ from the standard medium, the large potential(s) was abolished but not the earlier response (f). The early response (deflection) can now be clearly seen. C]: the recovery of the later potential 10 min after reintroduction of Ca 2+ into the standard medium. D, E: an example of LTP formation of a single slice when a tetanic stimulation at 50 Hz for 20 s was applied at 0 rain. Each trace in E is the average (Signal Processor NEC-Sanei 7T18) of I0 responses. Specimens in E correspond to the plots in D. The amplitude of PSP was measured from the base line of the potential to the peak of negativity (downward deflection). Note that there was no change in the amplitude of the earlier potential but only the PSP was enhanced. ON, optic nerve; SC, superior colliculus; IC, inferior colliculus; SG, superficial grey layer; OL, optic layer; IG, intermediate grey layer; IW, intermediate white layer; DG, deep grey layer; DW, deep white layer.

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MgSO4 1.3, CaCI2 2, and NaHCO3 26) equilibrated with 95% 02 and 5% CO2 (pH = 7.4). Thereafter each slice was transferred to a recording chamber under a stereomicroscope and submerged in the perfusion medium. Each layer of the SC can easily be visualized and identified (see Fig. 1B) under the stereomicroscope. The chamber was perfused continuously with the standard medium at a rate of 8 ml/min. The temperature in the chamber was kept at 35°C throughout the experiment. The arrangement of the stimulating and recording electrodes is also shown in Fig. lB. The evoked postsynaptic field potential (PSP) was recorded from the superficial grey layer (SGL) of the SC using a glass microelectrode containing 2 M NaCI. The stimulating electrodes were placed in the optic layer (OL) of the SC. Square pulse of 100 /ts in duration was used for stimulation (stimulator, Nihon Koden SEN 7103) which was at the rate of 0.5 Hz. By raising the strength of the stimulation, maximum amplitude was first noted and the stimulus intensity was adjusted to obtain a response which was about 1/3 of the maximum amplitude before the experiments were started. Tetanic stimulations at 50 Hz for 20 s were used in the experiment for the generation of LTP, because this parameter was found to be most effective for the induction of LTP. Electrical responses were recorded with an oscilloscope (Nihon Koden VCI0). O,L-APV (Sigma, St. Louis) at concentrations of 100 and 500 pM was prepared and dissolved in the standard medium before the experiment. Fig. I C shows that two kinds of potentials were recorded from this preparation after stimulation to the optic layer. The earlier potential was a short latency (less than 1.0 ms at peak latency) and low amplitude response. The later potential with a longer latency (2-4 msec at peak latency) was a high-amplitude response. The earlier potential was not blocked by the removal of Ca 2+ from the standard medium (see Fig. IC2). The later potential, however, was abolished by the removal of the Ca 2+ (see Fig. 1C2). It should be noted that the same pattern of responses were also recorded after stimulation to the optic nerve but could not be recorded in the slice from the animals which were enucleated contralaterally 12 days before the experiment. It is generally accepted [6, 13] that the neurons in the superficial grey layer harbor the terminals of the contralateral retinotectal fibers running through optic layer. Thus the above results suggest that the potential with longer latency and high amplitude was the monosynaptic PSP elicited by OL stimulation, and the response with short latency and low amplitude represented the presynaptic fiber potential or the antidromic activities of the SGL neurons. To determine the most effective parameters to form LTP, tetanic stimulation at 50 Hz, 100 Hz or 200 Hz, for 1, 5, 10, 20 or 30 s in duration (5 slices for each test) was applied to the OL. Among these combinations of frequency and duration of the tetanus, the stimulation of 50 Hz in frequency and 20 s in duration was found to be most effective for the induction of LTP. Fig. 1D, E shows an example of the formation of the LTP recorded from this preparation using the tetanic stimulation at 50 Hz and for 20 s. During application of 500 pM of D,L-APV, tetanic stimulation (50 Hz and 20 s) to the OL did not increase the amplitude of the response from that of the control level (Fig. 2C). However, when APV was removed from the medium, the amplitude

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Fig. 2. The formation of LTP in the superior colliculus and the effect of application of D,L-APV on LTP formation. Curve A indicates the time course of formation of LTP of the control slices in the standard perfusion medium without application of D,L-APV. Curves B and C indicate the time course of the LTP formation during and after removal of D,L-APV. tet indicates the beginning of the tetanic stimulation (50 Hz for 20 s) for A (tetA), B (tetB) and C (tetct, and tetc2). A horizontal stippled bar indicates the period of the application of D,L-APV for B (100 gM) and C (500 gM). Each plot is the mean amplitude of 5 slices. Vertical bar indicates S.D, Note that 500 gM but not 100 gM of O,L-APV masked the appearance of LTP formation and LTP was observed when standard medium was reintroduced.

of the PSP was increased to 150% of the control value in 15 min. When the second tetanic stimulation was applied after the removal of the APV, the amplitude of the PSP showed a further increase to 170% of the control value in 10 min. D,L-APV at 100/zM concentration did not inhibit the appearance of LTP. In fact, LTP formation was at 150% control value at the end of 20 min of recording (Fig. 2B). It should be noted however, that D,L-APV alone even at a dose of 500/tM did not alter the amplitude of the PSP without tetanic stimulation (Fig. 2). The application of y-DGG at a concentration of 1 mM also inhibited the appearance of LTP, but after the removal of the agent from the medium, the phenomenon of LTP appeared (not shown in the figure) as mentioned above in the case of APV. The finding that 500/tM D,L-APV inhibited the appearance of LTP in the SC suggests that the NMDA receptor is involved here. This result is consistent with other earlier findings on the role of the NMDA receptor in LTP formation in the hippocampus [7, 10, 14, 15], although the concentration of APV to inhibit the formation of LTP was 25-100 /2M in hippocampus which is lower concentration than that reported here. In our experiment, however, mixed D,L-APV was used and Col-

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lingridge et al. [3] reported that L-APV had a very weak inhibitory effect for the N M D A response compared with D-APV. To get the precise dose-response of the effect of APV on the inhibition of LTP formation requires further study using the pure optical isomers of D,L-APV. However, the inhibition of LTP appearance by large doses of D,L-APV (500/tM) and y-DGG (1 mM) suggest that the glutamate receptor, probably N M D A receptor, can be involved in the process of the appearance of LTP in SGL. Concerning the masking effect of APV on the appearance of LTP, it could be speculated that even during application of D,L-APV tetanic stimulation caused the modification of presynaptic plasticity for the induction of LTP such as the increase of release of the neurotransmitter from the presynaptic terminals or the induction of morphological change at the presynapse, even though the LTP phenomenon could not be observed because of the block of the N M D A receptor on the postsynaptic membrane, and that removal of the inhibition by the N M D A antagonist may allow the N M D A receptor to function and induce the LTP. Thus the presynaptic event may be essential for the initiation of LTP, if the N M D A receptor is located only on the postsynaptic membrane, although this possibility is to be studied further in the SC. It should be noted that once the LTP was formed, the increase in the response was not inhibited by the application of APV (500/tM) (not shown in the figure). This agrees with the hypothesis that N M D A channels of the postsynaptic site may be involved in the appearance of LTP but do not appear to contribute to the maintenance of LTP [4]. The same kind of masking effect for the LTP formation was also observed in the hippocampus in which y-DGG masked the appearance of LTP in perforant path [5]. LTP can be induced by a wide range of tetanus frequency, intensity and duration. As for the stimulation parameters, LTP was easily induced in the hippocampus by a tetanus applied for l s at 100 Hz [14] and a total of 10(~200 pulses were enough for inducing LTP. Furthermore, at lower frequencies of stimulation (below 50 Hz) a heterosynaptic response depression can be seen following the termination of tetanus [14]. Our experiments with the SC slice indicated that a tetanic frequency of 50 Hz and 20 s in duration was most effective for the LTP formation and a total of 1000 pulses were required for the formation of distinct LTP. In the goldfish optic tectum LTP was formed by the tetanus at 1-5 Hz for 20 s [9]. Thus the optimal stimulation parameters for the formation of LTP may differ by the region of the brain, by the efficacy of synapses or by species. Biochemical evidence from studies using the SC has shown that corticotectal fibers are glutamatergic and that while it has not been well documented, the presence of the glutamatergic innervation is suspected for the retinotectal pathway [6]. Our results show, however, that specific glutamatergic innervation might occur in the retinotectal pathway, because the N M D A receptor, a glutamatergic receptor, might be involved in the neurotransmission of this pathway. The finding that LTP can be reliably recorded in the SC is important and significant because it is the first demonstration of this phenomenon in the SC of the mammalian brain. The literature has shown that SC is important in the integration of the

113 a u d i t o r y , v i s u a l a n d s o m a t o s e n s o r y i n f o r m a t i o n [13, 16]. T h e r e f o r e , it is h i g h l y possible t h a t the L T P f o r m a t i o n h e r e m a y p l a y a n i m p o r t a n t r o l e in t h e g e n e r a l c o o r d i n a tion of the information from the retina, cortex and the brainstem, although the true m e c h a n i s m o f t h e L T P f o r m a t i o n in S C r e m a i n s to be s t u d i e d f u r t h e r . 1 Arakawa, T. and Okada, Y., Dual effect of ),-aminobutyric acid (GABA) on neurotransmission in the superior colliculus slices from the guinea pig, Proc. Jpn. Acad., 63 Ser. B (1987) 389-392. 2 Bliss, T.V.P. and Lomo, T.J., Long-lasting potentiation of synaptic transmission in the dentate area of the anesthetised rabbit following stimulation of the perforant path, J. Physiol. (Lond.), 232 (1973) 331-356. 3 Collingridge, G.L., Kehl, S.J. and McLennan, H., The antagonism of amino acid-induced excitations of rat hippocampal CA I neurones in vitro, J. Physiol. (Lond.), 334 (1983) 19-31. 4 Collingridge, G.L. and Bliss, T.V.P., NMDA receptors - their role in long-term potentiation, Trends Neurosci., l0 (1987) 288-293. 5 Dolphin, A.C., The excitatory amino-acid antagonist ),-o-glutamylglycine masks rather than prevents long term potentiation of the perforant path, Neuroscience, l0 0983) 377-383. 6 Fosse, V.M. and Fonnun, F., Effects of kainic acid and other excitotoxins in the rat superior colliculus: relations to glutamatergic afferents, Brain Res., 383 (1986) 28-37. 7 Harris, E.W., Ganong, A. and Cotman, C., Long-term potentiation in hippocampus involves activation of N-methyl-o-aspartate receptors, Brain Res., 323 (1984) 132-137. 8 Kawai, N. and Yamamoto, C., Effect of 5-hydroxytryptamine, LSD and related compounds on electrical activities evoked in vitro in thin sections from the superior colliculus, Int. J. Neuropharmacol., 8 (1969) 437~,49. 9 Lewis, D. and Teyler, T., Long-term potentiation in the goldfish optic tectum, Brain Res., 375 (1986) 246-250. l0 Morris, R.G.M., Anderson, E., Lynch, G.S. and Baudry, M., Selective impairment of learning and blockade of long-term potentiation by an N-methyl-D-aspartate receptor antagonist, AP5, Nature (Lond.), 319 (1986) 774-776. I 10kada, Y., Distribution of 3,-aminobutyric acid (GABA) in the layers of superior colliculus of the rabbit, Brain Res., 75 (1974) 362-365. 12 Okada, Y. and Saito, M., Inhibitory action of adenosine, 5-HT (serotonin) and GABA (3,-aminobutyric acid) on the postsynaptic potential (PSP) of slices from olfactory cortex and superior colliculus in correlation to the level of cyclic AMP, Brain Res., 160 (1979) 368 371. 13 Sprague, J.M., Mammalian tectum: intrinsic organization, afferent inputs, and integrative mechanisms, Neurosci. Res. Prog. Bull., 13 (1975) 204-213. 14 Teyler, T.J. and DiScenna, P., Long-term potentiation, Annu. Rev. Neurosci., 10 (I 987) 131- 161. 15 Wigstr6m, H. and Gustafsson, B., A possible correlate of the synaptic condition for long-lasting potentiation in the guinea pig hippocampus in vitro, Neurosci. Lett., 44 (1984) 327-332. 16 Wurtz, R.H. and Albano, J.E., Visual motor function of the primate superior colliculus, Annu. Rev. Neurosci., 3 (1980) 189-226.