Zhang Yan, Dixon Christine L, Keramidas Angelo, Lynch Joseph W
Queensland Brain Institute, University of Queensland, Brisbane, QLD 4072, Australia.
Neuropharmacology. 2015 Feb;89:391-7. doi: 10.1016/j.neuropharm.2014.10.026.
Glycine receptor (GlyR) chloride channels mediate fast inhibitory neurotransmission in the spinal cord and brainstem. Four GlyR subunits (α1-3, β) have been identified in humans, and their differential anatomical distributions underlie a diversity of synaptic isoforms with unique physiological and pharmacological properties. To improve our understanding of these properties, we induced the formation of recombinant synapses between cultured spinal neurons and HEK293 cells expressing GlyR subunits of interest plus the synapse-promoting molecule, neuroligin-2A. In the heterosynapses thus formed, recombinant α1β and α3β GlyRs mediated fast decaying inhibitory postsynaptic currents (IPSCs) whereas α2β GlyRs mediated slow decaying IPSCs. These results are consistent with the fragmentary information available from native synapses and single channel kinetic studies. As β subunit incorporation is considered essential for localizing GlyRs at the synapse, we were surprised that α1-3 homomers supported robust IPSCs with β subunit incorporation accelerating IPSC rise and decay times in α2β and α3β heteromers only. Finally, heterosynapses incorporating α1(D80A)β and α1(A52S)β GlyRs exhibited accelerated IPSC decay rates closely resembling those recorded in native synapses from mutant mice homozygous for these mutations, providing an additional validation of our technique. Glycinergic heterosynapses should prove useful for evaluating the effects of drugs, hereditary disease mutations or other interventions on defined GlyR subunit combinations under realistic synaptic activation conditions.
甘氨酸受体（GlyR）氯离子通道介导脊髓和脑干中的快速抑制性神经传递。在人类中已鉴定出四种甘氨酸受体亚基（α1 - 3，β），它们不同的解剖分布构成了具有独特生理和药理特性的多种突触亚型的基础。为了更好地理解这些特性，我们诱导培养的脊髓神经元与表达感兴趣的甘氨酸受体亚基以及突触促进分子神经连接蛋白 - 2A的HEK293细胞之间形成重组突触。在如此形成的异突触中，重组α1β和α3β甘氨酸受体介导快速衰减的抑制性突触后电流（IPSC），而α2β甘氨酸受体介导缓慢衰减的IPSC。这些结果与来自天然突触和单通道动力学研究的零碎信息一致。由于β亚基的掺入被认为是将甘氨酸受体定位在突触处所必需的，我们惊讶地发现α1 - 3同聚体支持强大的IPSC，而β亚基的掺入仅加速了α2β和α3β异聚体中IPSC的上升和衰减时间。最后，包含α1（D80A）β和α1（A52S）β甘氨酸受体的异突触表现出加速的IPSC衰减率，与这些突变纯合的突变小鼠天然突触中记录的衰减率非常相似，这为我们的技术提供了额外的验证。甘氨酸能异突触应该被证明对评估药物、遗传性疾病突变或其他干预措施在现实突触激活条件下对特定甘氨酸受体亚基组合的影响是有用的。
