Zdilar D, Luntz-Leybman V, Frostholm A, Rotter A
Department of Pharmacology, Ohio State University, Columbus 43210.
J Comp Neurol. 1992 Dec 22;326(4):580-94. doi: 10.1002/cne.903260407.
Gamma aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the mammalian cerebellum. Cerebellar granule, Purkinje, and deep nuclear neurons are known to receive GABAergic afferents. Since GABA exerts its inhibitory effects via GABA receptors, it is of interest to determine the temporal relationship between the formation of GABAergic synapses and the expression of genes coding for the GABA receptor. In a previous study, we have examined the developmental expression of binding sites for [3H]muscimol, which binds with high affinity to the beta subunits of the GABAA/benzodiazepine (GABAA/BZ) receptor. In the present study, [35S]cRNA probes were used to examine the appearance and distribution of GABAA/BZ beta 1, beta 2, and beta 3 subunit mRNAs in the developing C57BL/6 mouse cerebellum by in situ hybridization. In the adult cerebellum, the distribution of the three subunit mRNAs was clearly different, despite considerable overlap, and their temporal expression differed throughout postnatal development. The beta 1 hybridization signal appeared within the cerebellar cortex during the second postnatal week as a discrete band at the interface of the molecular and granule cell layers. Grains were distributed diffusely over small densely staining cells surrounding the Purkinje cells; relatively few grains were visible over Purkinje cell bodies themselves. This distribution may reflect an association with Bergmann glia or basket cells. The beta 2 and beta 3 hybridization signals were present considerably earlier than that of the beta 1 mRNA. The beta 2 signal was present at birth in the molecular/Purkinje cell layer; as development progressed, the signal became increasingly intense over both granule and Purkinje cells. At birth, the beta 3 subunit mRNA was present in the external germinal and molecular layers, later becoming largely localized within the granule cell layer. Dense beta 2 and beta 3 cRNA probe labeling was present over the adult granule cell layer. Moderate levels of beta 2 signal were seen over Purkinje cell bodies; considerably less labeling was observed with the beta 3 probe. The adult distribution of beta 2 and beta 3 cRNA probes showed good spatial correspondence with the known GABAA receptor beta subunit markers, [3H]-muscimol and the mAb 62-3G1 antibody, each being present within the granule cell layer. Our results indicate that the temporal expression of GABAA/BZ receptor beta subunit messages within a given cell type may be independently regulated, and that acquisition of the beta 2 and beta 3 mRNAs occurs before these cells become integrated into mature synaptic circuits.
γ-氨基丁酸（GABA）是哺乳动物小脑中主要的抑制性神经递质。已知小脑颗粒细胞、浦肯野细胞和深部核神经元接受GABA能传入纤维。由于GABA通过GABA受体发挥其抑制作用，因此确定GABA能突触的形成与编码GABA受体的基因表达之间的时间关系很有意义。在先前的一项研究中，我们检测了与GABAA/苯二氮䓬（GABAA/BZ）受体的β亚基具有高亲和力结合的[3H]蝇蕈醇结合位点的发育表达。在本研究中，使用[35S]cRNA探针通过原位杂交检测发育中的C57BL/6小鼠小脑中GABAA/BZβ1、β2和β3亚基mRNA的出现和分布。在成年小脑中，尽管有相当大的重叠，但三种亚基mRNA的分布明显不同，并且它们在出生后的整个发育过程中的时间表达也不同。β1杂交信号在出生后第二周出现在小脑皮质内，在分子层和颗粒细胞层的界面处呈离散带。银粒弥漫分布在浦肯野细胞周围的小的密集染色细胞上；在浦肯野细胞体本身可见的银粒相对较少。这种分布可能反映了与伯格曼胶质细胞或篮状细胞的关联。β2和β3杂交信号比β1 mRNA出现得早得多。β2信号在出生时存在于分子层/浦肯野细胞层；随着发育的进行，颗粒细胞和浦肯野细胞上的信号变得越来越强。出生时，β3亚基mRNA存在于外生发层和分子层，后来主要定位于颗粒细胞层。成年颗粒细胞层上存在密集的β2和β3 cRNA探针标记。在浦肯野细胞体上可见中等水平的β2信号；用β3探针观察到的标记明显较少。β2和β3 cRNA探针的成年分布与已知的GABAA受体β亚基标记物[3H] - 蝇蕈醇和单克隆抗体62 - 3G1抗体显示出良好的空间对应关系，它们都存在于颗粒细胞层内。我们的结果表明，给定细胞类型内GABAA/BZ受体β亚基信息的时间表达可能受到独立调节，并且β2和β3 mRNA的获得发生在这些细胞整合到成熟突触回路之前。
