REGULATION OF INTRACELLULAR CL- LEVELS BY NA-DEPENDENT CL- COTRANSPORT DISTINGUISHES DEPOLARIZING FROM HYPERPOLARIZING GABA(A) RECEPTOR-MEDIATED RESPONSES IN SPINAL NEURONS()

Rohon-Beard (RB) spinal neurons of Xenopus larvae are depolarized by G ABA. To study the mechanisms underlying this distinctive response, int racellular and patch-clamp recordings were made from RB neurons in sit u. The intracellularly recorded GABA reversal potential (E(Rev)) was n ear -30 mV in normal saline and was similar to 25 mV more negative in Na+-free saline. Whole-cell recordings from RB neurons and from neighb oring dorsolateral interneurons (DLi) revealed that GABA responses of both cells were mediated by GABA(A) receptors. Currents elicited by GA BA were mimicked by muscimol and reversibly blocked by bicuculline, an d E(Rev) shifted with changes in Cl- concentration ([Cl-]) in agreemen t with Cl- selectivity. In perforated patch recordings, E(Rev) for RB cells was significantly more positive than for DLi cells (-38 vs -63 m V), indicating that intact RB cells maintain higher levels of intracel lular Cl-. Replacement of external Na+ or exposure to the Cl- transpor t inhibitor bumetanide (100 mu M) shifted RB cell E(Rev) to more negat ive values, consistent with Na+-dependent Cl- cotransport contributing to higher internal [Cl-]. In contrast, these treatments did not chang e DLi cell E(Rev). The results indicate that a Na+-dependent Cl- trans port mechanism underlies GABA(A) receptor-mediated depolarizing Cl- co nductances in RB neurons. Thus, both inhibitory and excitatory GABA re sponses appear to be present during the same developmental period in v ivo. GABA may stimulate Ca2+ influx in RB neurons because the intracel lular GABA E(Rev) is above the threshold for low voltage-activated Ca2 + channels.