不同刺激形式在成年大鼠海马CA1区诱导长时程压抑的谷氨酸受体机制_英文_

不同刺激形式在成年大鼠海马CA1区诱导长时程压抑的谷氨酸受体机制_英文_ Acta Physiologica Sinica, April 25, 2008, 60 (2): 270-274 270 Different glutamate receptor mechanisms in long-term depression induced by different stimulus patterns in the CA1 area of adult rat hippocampus *CHEN Li, HAN Tai-Zhen, JIANG Ma-Li Department of Physiology and Pathophysiology, School of Medicine, Xi’an Jiaotong University, Xi’an 710061, China Abstract: Previous reports suggested that a novel stimulus pattern of multi-train stimulus at low-frequency (2-Hz or 5-Hz) could induce stable long-term depression (LTD) in the CA1 area of adult rat hippocampus. In the present study, in order to determine the mechanism in LTD induced by the two novel tetanus patterns, changes in the population spikes (PS) in the hippocampal CA1 area of adult rats following the multi-train stimulus in the presence of AP5 [antagonist of N-methyl-D-aspartate receptors (NMDARs)] or MCPG [antagonist of type I/II metabotropic glutamate receptors (mGluRs)] were recorded. The results showed that both AP5 and MCPG inhibited the LTD induced by 2-Hz multi-train stimulus. The mean amplitude of population spikes (PSA) normalized to the baseline was (96.0?3.5)% after applying AP5 (n=10) and (95.7?4.1)% after applying MCPG (n=8), respectively, measured at 20 min post-tetanus. While 5-Hz multi-train tetanus failed to induce LTD in the presence of MCPG. The mean PSA was (73.6?4.4)% (n=10) and (98.2?8.9)% (n=8) in the presence of AP5 and MCPG, respectively, measured at 35 min post-tetanus. So it is suggested that LTD induced by 2-Hz multi-train tetanus involves co-activation of NMDARs and mGluRs, while LTD induced by 5-Hz multi-train tetanus is only related to activation of mGluRs. Key words: long-term depression; hippocampus; N-methyl-D-aspartate receptors; metabotropic glutamate receptors 不同刺激形式在成年大鼠海马 CA 1 区诱导长时程压抑的谷氨酸受体机制 *陈 丽,韩太真,蒋马莉 西安交通大学医学院生理与病理生理学系，西安 710061 摘 要:前期研究显示低频率多串刺激能够在成年大鼠海马 CA1 区诱发稳定的长时程压抑(long-term depression, LTD)，而这种 LT D 的受体机制目前还不清楚。本研究采用成年大鼠海马脑片标本，电刺激 Sch affer 侧枝传入纤维，在 CA1 区锥体细胞 层 记录 混凝土 养护记录下载土方回填监理旁站记录免费下载集备记录下载集备记录下载集备记录下载 群体锋电位(population spikes, PS)，并分别应用 N - 甲基 -D - 天冬氨酸(N -methyl-D-aspartate, NMDA)受体和代谢型谷氨 酸(metabotropic glutamate, mGlu)受体的拮抗剂 AP5 和 MCPG，观察两组低频率(2-Hz 和 5-H z)多串刺激能否诱导 LT D，以揭 示不同刺激形式诱导成年大鼠 LT D 的可能受体机制。结果显示，AP 5 和 MC P G 都能抑制由 2-H z 多串刺激诱导的 LT D :强 直刺激后 20 min 时 PS 幅度分别为基础值的(96.0?3.5)% (n=10)和(95.7?4.1)% (n=8 )。MCPG 能够抑制 5-H z 多串刺激诱导的 LTD 的产生，而 AP5 不能:分别应用 AP5 和 MCPG 后，强直刺激后 35 min 时 PS 的幅度分别为基础值的(73.6?4.4)% (n=10) 和(9 8.2?8 .9)% (n =8)。以上结果提示，2-Hz 多串刺激诱导的 LT D 可能依赖于 NMDA 受体与 mGlu 受体的共同活化，而 5-Hz 多串刺激诱导的 LTD 只与 mGlu 受体有关。因此，不同频率的多串刺激诱导的 LT D 涉及不同的谷氨酸受体机制。 关键词:长时程压抑;海马;NMDA 受体;代谢型谷氨酸受体中图分类号:Q4 2 ;R33 8 . 8 Received 2007-09-12 Accepted 2007-12-27 This work was supported by the National Natural Science Foundation of China (No. 30170310). *Corresponding author. Tel: +86-29-82655274; Fax: +86-29-82655274; E-mail: htzhen@mail.xjtu.edu.cn Glutamate is the main excitatory neurotransmitter in the brain. Its receptors can be divided into ionotropic and metabotropic receptors, and they have an important role [1]in learning and memory. Early studies in the CA1 area of hippocampus showed that long-term depression (LTD) was dependent on the activation of N-methyl-D-aspar- tate (NMDA) receptors (NMDARs). Subsequently, more animal experiments showed that LTD may also be de- pendent on the activation of metabotropic glutamate re- [2]ceptors (mGluRs) in this area. These results suggest that perhaps there are two di sti nct for ms of LTD [3](NMDAR-LTD and mGluR-LTD) in the CA1 area. Low- frequency stimulus (LFS: 1 Hz, 15 min) was able to in- duce NMDAR-LTD in the CA1 area of hippocampus in [4]Fig. 1. LTD induced by two multi-train stimuli at low-frequency. young animals (< 40 d), and prolonged LFS (1 Hz, 30 The protocol 1 (c, n=10) was 600 pulses at 2 Hz divided into 5 trains min) or paired-pulse LFS (PP-LFS: 900 paired stimuli) with 60-s inter-train interval, and the protocol 2 (b, n=8) was 600 with 200-ms paired-pulse interval (PPI) could induce pulses at 5 Hz divided into 5 trains with 96-s inter-train interval. a, [5]NMDAR-LTD in adult animal hippocampus. NMDAR- basline. PSA, the mean amplitude of population spikes. LTD could also be induced in young animal hippocam- pus (< 50 d) by altering the PP-LFS protocol (50-ms PPI). In contrast, the same stimulus protocol induced 1 MATERIALS AND METHODS [5]mGluR-LTD in adult animals (12-16 weeks). In addition, [6] [7] 1.1 Animals when the NMDARsand mGluRswere separately ac- tivated by pharmacological method, LTD can also be Transverse hippocampal slices (400 μm) were prepared induced. In conclusion, different factors, such as animal from male adult Sprague-Dawley rats weighing 180-250 g. age and stimulus patterns, may have influence on the Experiments were conducted in compliance with the labo- mechanisms in LTD induction. Namely, depending on ratory animal protocol utilized in the Medical College of the experimental conditions, either NMDAR-LTD or Xi’an Jiaotong University. mGluR-LTD could be induced in the same CA1 pyrami- 1.2 Slice preparation dal cells. However, these two distinct forms of LTD are Rat was decapitated, and the hippocampus was quickly likely to be related to different signal transduction cas- removed and cut into 400-μm thick slices. The slices cades and expression mechanisms, even though they exist were placed on a nylon mesh, incubated in an interface [3]in the same pyramidal cells. recording chamber maintained at (31?1) ?C and permit- We previously reported that a novel stimulus pattern ted to recover for at least 1.5 h before recording. Some of multi-train stimulus at low frequency could induce a slices were incubated in artificial cerebrospinal fluid [8]stable LTD in the CA1 area of adult rat hippocampus. (ACSF, control), others in ACSF with 50 μmol/L AP5 Two multi-train stimuli were adopted: 600 pulses at 2 Hz (NMDAR antagonist) or 500 μmol/L MCPG (type I/II divided into 5 trains with 60-s inter-train interval, and mGluR antagonist), respectively. ACSF contained (in 600 pulses at 5 Hz divided into 5 trains with 96-s inter- mmol/L) NaCl 124, KCl 5, NaHCO26, MgSO1.3, 3 4 train interval. The results showed that LTD produced by [9] KHPO1.2, CaCl2.4, glucose 10 . ACSF was satu- 24 2 different stimulus frequencies with the same total pulses rated with 95% Oand 5% CO 2 2possesses distinct properties of latency (the period from 1.3 Electrophysiology the end of tetanus to a beginning of LTD) and amplitude (Fig.1). Based on the results described above, we assume Population spikes (PS) were recorded in the pyramidal that two different receptor mechanisms may be involved cell layer of the CA1 area using micropipettes filled with in the LTD produced by these two distinct stimulus 2 mol/L NaCl (impedances of 1-3 MΩ), following patterns. stimulation of the Schaffer collateral-commissural fibers. Acta Physiologica Sinica, April 25, 2008, 60 (2): 270-274 272 The stimulus intensity was two folds of the threshold (n=10); but failed to induce LTD in the presence of MCPG: intensity with 0.1-ms duration. The strength of synap- the mean PSA normalized to the baseline was (98.2?8.9)% tic response was quantified by measuring the peak-to- and (93.2?3.1)% measured at 35 min and 80 min post- peak amplitude of population spikes (PSA). Stable tetanus, respectively (n=8) (Fig.2B). In addition, ampli- baseline responses were recorded for at least 40 min tudes of LTD induced by protocol 2 with [(72.1?4.1)%, with test stimuli at 0.017 Hz only in those slices that n=10] or without [(67.7?3.4)%, n=8] treatment of AP5 showed a stable slope and amplitude. After obtaining a showed no significant difference at 80 min post-tetanus stable baseline recording, we applied two protocols of (Fig.3). tetanus at the test intensity to induce LTD as previously: protocol 1 was 600 pulses at 2 Hz divided into 5 trains with 60-s inter-train interval, and the protocol 2 was 600 pulses at 5 Hz divided into 5 trains with 96-s inter- [8]train interval. The responses after tetanus were re- corded for more than 90 min. The standard of LTD formation indicates that the PSA after tetanus decreased by 20% or more compared t o that i n the ba seli ne recording, and this depression lasted for more than 30 min. At the end of the LTD recording, we tested the hippocampal slices by applying a single tetanus train at high-frequency stimulus (HFS: 100 Hz, 1 s; 30-60 μA, 0.1-ms duration) to the Schaffer collateral fibers. 1.4 Data analysis Data were expressed as means?SEM. Statistical analysis was performed by two-tailed independent Student t-test. P<0.05 were considered to be statistically significant. 2 RESULTS Control stimulus in protocol 1 (control 1, 5 trains at 2 Hz), induced LTD within 15-25 min after tetanus: the mean PSA normalized to the baseline was (78.1?3.1)% and (57.5?2.8)% measured at 20 min and 80 min post-tetanus, respectively (n=10). However, protocol 1 stimulus, given in the presence of AP5 (50 μmol/L) or MCPG (500 μmol/L), failed to bring the obvious changes in PSA (Fig.2A): the mean PSA normalized to the baseline measured at 20 min and 80 min post-tetanus in the presence of AP5 was (96.0?3.5)% and (100.0?3.1)%, respectively (n=10); that in the presence of MCPG was (95.7?4.1)% and (94.5?4.1)%, respectively (n=8). Control stimulus in protocol 2 (control 2, 5 trains at 5 Hz) could also induce stable LTD within 30-40 min after teta- nus in the hippocampal slices bathed in the ACSF: the mean Fig. 2. Changes in the mean amplitude of population spikes (PSA) in PSA normalized to the baseline was (78.3?5.3)% and hippocampal slices treated with AP5, MCPG after 2-Hz and 5-Hz (67.7?3.4)% measured at 35 min and 80 min post-tetanus, multi-train tetanus. A: 2-Hz multi-train tetanus did not induce LTD respectively (n=8). Nevertheless, protocol 2 stimulus still in the presence of AP5 or MCPG. B: The induction of LTD by 5-Hz induced LTD in the presence of AP5: the mean PSA nor- multi-train tetanus was unaffected in the presence of AP5, but was blocked in the presence of MCPG. a, basline; b, AP5; c, MCPG; d, malized to the baseline was (73.6?4.4)% and (72.1?4.1)% control. measured at 35 min and 80 min post-tetanus, respectively quired for LTD induction. In addition, in CA1 area, LTD induced by DHPG (type I mGluR agonist) results from modulation of the presynaptic calcium currents result- 2+ [14]ing in Cainflux mediated by type I mGluR, and that 2+ might require the NMDAR activation to increase the Ca [15]influx. The present study showed that the induction of LTD by 2-Hz multi-train stimulus required the co-activa- tion of NMDARs and mGluRs, suggesting that this LTD 2+ induction might require more Caso as to reach the proper 2+level of membrane potential and [Ca]. Whereas the for- iFig. 3. Comparison of the mean amplitude of population spikes mation of LTD induced by 5-Hz multi-train stimulus might (PSA) normalized to the baseline in LTD induced by 5 Hz multi- 2+require lower [Ca]. Since MCPG is an antagonist of type train tetanus with or without AP5 treatment at 80 min post-tetanus. i I/II mGluRs, more studies are needed to assess whether There was no significant difference between them. both type I and II mGluRs or only one of them participate in the LTD induction. 3 DISCUSSION REFERENCES The data presented here confirm that multi-train stimulus patterns at low-frequency were able to induce a stable LTD Watkins JC, Evans RH. Excitatory amino acid transmitters. Annu 1 via different receptor mechanisms. LTD induced by 2-Hz Rev Pharmacol Toxicol 1981; 21: 165-204. multi-train tetanus needed co-activation of NMDARs and Bolshakov VY, Siegelbaum SA. 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