Effect of agonist concentration on the lifetime of GABA-activated membrane channels in spinal cord neurons.

Abstract

The inhibitory neurotransmitter gamma-aminobutyric acid (GABA) hyperpolarizes spinal neurons by activating bicuculline sensitive GABAA receptors coupled to chloride permeable ionic channels in the cell membrane. Single channel measurements using outside-out membrane patches from cultured mouse spinal neurons showed that there is a large excess of brief openings of these channels, compared to the number predicted from a simple, two-state model of channel function. The cumulative open time distributions for channel openings were well fit by the sum of two exponential terms, a fast component with amplitude Nf and time constant TAUf, and a slow term with amplitude Ns and time constant TAUs. These kinetics could arise if monoliganded GABAA receptors triggered an open state less stable than that induced by the biliganded receptor. A prediction of this hypothesis is that the ratio Nf/Ns should decline in an approximately linear fashion with increasing doses of GABA. This was found to be the case, as Nf/Ns declined by a factor of 4.8 on increasing the GABA dose from 0.5 microM to 5 microM. Application of 1.25 microM GABA produced an intermediate value of Nf/Ns, as predicted. In contrast the parameters TAUf and TAUs were not significantly influenced by GABA concentration. Spontaneous, bicuculline sensitive currents were seen in some patches. These events resembled currents triggered by low doses of exogenous GABA, with regard to the values of Nf/Ns, TAUf and TAUs. These results suggest that the spontaneous currents are triggered by endogenous GABA molecules, present in the culture environment at a concentration of 0.5 microM or less.