Apamin as a selective blocker of the calcium-dependent potassium channel in neuroblastoma cells: voltage-clamp and biochemical characterization of the toxin receptor.

Abstract

The interaction of apamin, a bee venom neurotoxin, with mouse neuroblastoma cell membrane was described. Voltage-clamp analyses showed that apamin at low concentrations specifically blocked the Ca2+-dependent K+ channel in differentiated neuroblastoma cells. Binding experiments with highly radiolabeled toxin indicated that the Kd of the apamin-receptor complex in differentiated neuroblastoma cells was 15-22 pM and the maximal binding capacity was 12 fmol[femtomol]/mg protein. The receptor was destroyed by proteases, suggesting that it was a protein. The binding capacity of neuroblastoma cells for 125I-apamin dramatically increased during the transition from the nondifferentiated to the differentiated state. The number of Ca2+-dependent K+ channels was at most 1/5 the number of fast Na+ channels in differentiated neuroblastoma. The binding of 125I-apamin to its receptor was antagonized by monovalent and divalent cations. Na+ inhibition of the binding of 125I-apamin was of the competitive type (Kd(Na+) = 44 mM). Guanidinium and guanidinated compounds such as amiloride or neurotensin prevented binding of 125I-apamin, the best antagonist being neurotensin.