Kim H S, Choi H S, Lee S Y, Oh S
College of Pharmacy, Chungbuk National University, Cheongju, 361-763, Chungbuk, South Korea.
Brain Res. 2000 Oct 13;880(1-2):28-37. doi: 10.1016/s0006-8993(00)02687-1.
In the present study, we have investigated the effects of prolonged inhibition of NMDA receptor by infusion of subtoxic dose of MK-801 to examine the modulation of GABA(A) receptor binding and GABA(A) receptor subunit mRNA level in rat brain. It has been reported that NMDA-selective glutamate receptor stimulation alters GABA(A) receptor pharmacology in cerebellar granule neurons in vitro by altering the levels of selective subunit. However, we have investigated the effect of NMDA antagonist, MK-801, on GABA(A) receptor binding characteristics in discrete brain regions by using autoradiographic and in situ hybridization techniques. The GABA(A) receptor bindings were analyzed by quantitative autoradiography using [3H]muscimol, [3H]flunitrazepam, and [35S]TBPS in rat brain slices. Rats were infused with MK-801 (1 pmol/10 microl per h, i.c.v.) for 7 days, through pre-implanted cannula by osmotic minipumps (Alzet, model 2 ML). The levels of [3H]muscimol binding were highly elevated in almost all of brain regions including cortex, caudate putamen, thalamus, hippocampus, and cerebellum. However, the [3H]flunitrazepam binding and [35S]TBPS binding were increased only in specific regions; the former level was increased in parts of the cortex, thalamus, and hippocampus, while the latter binding sites were only slightly elevated in parts of thalamus. The levels of beta2-subunit were elevated in the frontal cortex, thalamus, hippocampus, brainstem, and cerebellar granule layers while the levels of beta3-subunit were significantly decreased in the cortex, hippocampus, and cerebellar granule layers in MK-801-infused rats. The levels of alpha6- and delta-subunits, which are highly localized in the cerebellum, were increased in the cerebellar granule layer after MK-801 treatment. These results show that the prolonged suppression of NMDA receptor function by MK-801-infusion strongly elevates [3H]muscimol binding throughout the brain, increases regional [3H]flunitrazepam and [35S]TBPS binding, and alters GABA(A) receptor subunit mRNA levels in different directions. The chronic MK-801 treatment has differential effect on various GABA(A) receptor subunits, which suggests involvement of differential regulatory mechanisms in interaction of NMDA receptor with the GABA receptors.
在本研究中，我们通过注入亚毒性剂量的MK-801来研究长时间抑制NMDA受体的作用，以检测大鼠脑中GABA(A)受体结合及GABA(A)受体亚基mRNA水平的调节情况。据报道，NMDA选择性谷氨酸受体刺激可通过改变选择性亚基的水平，在体外改变小脑颗粒神经元中GABA(A)受体的药理学特性。然而，我们利用放射自显影和原位杂交技术，研究了NMDA拮抗剂MK-801对离散脑区中GABA(A)受体结合特性的影响。在大鼠脑片中，使用[3H]蝇蕈醇、[3H]氟硝西泮和[35S]TBPS，通过定量放射自显影分析GABA(A)受体结合情况。通过渗透微型泵（Alzet，型号2ML），经预先植入的套管，以1 pmol/10微升每小时的剂量向大鼠脑室内注入MK-801，持续7天。几乎在所有脑区，包括皮质、尾状壳核、丘脑、海马和小脑，[3H]蝇蕈醇结合水平都显著升高。然而，[3H]氟硝西泮结合和[35S]TBPS结合仅在特定区域增加；前者在部分皮质、丘脑和海马中升高，而后者的结合位点仅在部分丘脑中略有升高。在注入MK-801的大鼠中，额叶皮质、丘脑、海马、脑干和小脑颗粒层中β2亚基水平升高，而皮质、海马和小脑颗粒层中β3亚基水平显著降低。在MK-801处理后，高度定位于小脑中的α6和δ亚基水平在小脑颗粒层中升高。这些结果表明，通过注入MK-801对NMDA受体功能进行长时间抑制，可使全脑的[3H]蝇蕈醇结合显著升高，增加区域[3H]氟硝西泮和[35S]TBPS结合，并使GABA(A)受体亚基mRNA水平在不同方向上发生改变。慢性MK-801处理对各种GABA(A)受体亚基有不同影响，这表明在NMDA受体与GABA受体相互作用中存在不同的调节机制。
