Role of Cl–Co-Transporters in the Excitation Produced by GABAAReceptors in Juvenile Bovine Adrenal Chromaffin Cells

Abstract

GABA is the primary inhibitory neurotransmitter in the adult mammalian brain. However, in neonatal animals, activation of Cl–-permeable GABA receptors is excitatory and appears to depend on the expression of a Na+-K+-2Cl–cotransporter (NKCC) that elevates intracellular Cl–levels, leading to a depolarized Cl–equilibrium potential ( ECl). The change from excitation to inhibition appears to involve the expression of the K+/Cl–co-transporter, KCC2, which lowers intracellular Cl–levels resulting in a hyperpolarized ECl. In this study, we show that bovine chromaffin cells from 4- to 5-mo-old animals are excited by GABA. Activation of GABAAreceptors depolarizes the cells, opens voltage-dependent Ca2+channels, elevates [Ca2+]i, and promotes the release of catecholamines. Blockade of voltage-dependent Ca2+channels prevents the elevation of [Ca2+]iby GABA. The extrapolated anion reversal potential in these cells is approximately –28 mV, indicating a resting intracellular anion concentration of approximately 50 mM. Expression of KCC2 protein was not detected in the juvenile chromaffin cells. In contrast, clear expression of NKCC1 was observed. Blockade of NKCC1 should reduce the intracellular Cl–concentration and hyperpolarize ECl. Bumetanide, an NKCC1 blocker, reduced the elevation of [Ca2+]iby GABA. In some cells, activation of GABAAreceptors inhibits responses to excitatory neurotransmitters, even though GABA itself is depolarizing. Co-activation of cholinergic and GABAAreceptors in chromaffin cells produced elevations in [Ca2+]ithat were comparable to those produced by cholinergic receptors alone. Our data showing the selective expression of chloride co-transporters and the resulting strongly depolarized anion reversal potential may help explain how activation of GABAAreceptors causes sufficient excitation to elicit catecholamine release from chromaffin cells.