Angiotensin II regulates neuronal excitability via phosphatidylinositol 4,5-bisphosphate-dependent modulation of Kv7 (M-type) K~+ channels

摘要

Voltage-gated Kv7 (KCNQ) channels underlie important K~+ currents in many different types of cells, including the neuronal M current, which is thought to be modulated by muscarinic stimulation via depletion of membrane phosphatidylinositol 4,5-bisphosphate (PIP2)- We studied the role of modulation by angiotensin II (angioll) of M current in controlling discharge properties of superior cervical ganglion (SCG) sympathetic neurons and the mechanism of action of angioll on cloned Kv7 channels in a heterologous expression system. In SCG neurons, which endogenously express angioll ATI receptors, application of angioll for 2 min produced an increase in neuronal excitability and a decrease in spike-frequency adaptation that partially returned to control values after 10 min of angioll exposure. The increase in excitability could be simulated in a computational model by varying only the amount of M current. Using Chinese hamster ovary (CHO) cells expressing cloned Kv7.2 + 7.3 heteromultimers and ATI receptors studied under perforated patch clamp, angioll induced a strong suppression of the Kv7.2/7.3 current that returned to near baseline within 10 min of stimulation. The suppression was blocked by the phospholipase C inhibitor edelf osine. Under whole-cell clamp, angioll moder ately suppressed the Kv7.2/7.3 current whether or not intracellular Ca~2+ was clamped or Ca~2+ stores depleted. Co-expression of PI(4)5-kinase in these cells sharply reduced angioll inhibition, but did not augment current amplitudes, whereas co-expression of a PIP_2 5'-phosphatase sharply reduced current amplitudes, and also blunted the inhibition. The rebound of the current seen in perforated-patch recordings was blocked by the PI4-kinase inhibitor, wortmannin (50 /XM), suggesting that PIP_2 re-synthesis is required for current recovery. High-performance liquid chr omatographic analysis of anionic phospholipids in CHO cells stably expressing AT 1 receptors revealed that PIP_2 and phosphatidylinositol 4-phosphate levels are to be strongly depleted after 2 min of stimulation with angioll, with a partial rebound after 10 min. The results of this study establish how angioll modulates M channels, which in turn affects the integrative properties of SCG neurons.