Internal perfusion of isolated CNS neurons and the analysis of neuronal receptor potential by concentration-clamp technique.

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Electrophysiological studies have been performed on acutely isolated peripheral neurons from adult invertebrate and vertebrate preparations, heart muscles and smooth muscles. Investigations on single CNS neurons have been performed using cultured cells from fetal rats and mice. Other studies have involved the use of hippocampal pyramidal neurons dissociated acutely from both infant rats and adult guinea pogs. However, the neurons isolated by previous techniques lacked the distal dendric structures, and the original shape of soma was lost. Thus, a technique for isolating CNS neurons with well-preserved morphological structures has not been established. Here we have succeeded in isolating CNS neurons with a well-preserved morphological structure. The physiological functions (e.g., voltage-dependent and drug-gated chammels) in these isolated neurons were also well preserved. For example, isolated pyramidal cells from young and adult rat hippocampus were sensitive to all excitatory amino acids (glutamate, kainate, quisqualate and NMDA) and inhibitory amino acids (glycine, taurine, GABA). The NMDA response was dramatically potentiated by adding glycine at threshold concentration (10^<-6>M). In addition, the ionic selectivity of voltage-dependent Na channel of isolated CNS neurons was in the order of Li > Na > hydrazine > formamidine > guanidine > methylguanidine > monomethylamine. The order was quite similar to that in heart ventricle cell and squid giant axon.