Calcium affects OX1 orexin (hypocretin) receptor responses by modifying both orexin binding and the signal transduction machinery.
BRITISH JOURNAL OF PHARMACOLOGY(2014)
摘要
Background and PurposeOne of the major responses upon orexin receptor activation is Ca2+ influx, and this influx seems to amplify the other responses mediated by orexin receptors. However, the reduction in Ca2+, often used to assess the importance of Ca2+ influx, might affect other properties, like ligand-receptor interactions, as suggested for some GPCR systems. Hence, we investigated the role of the ligand-receptor interaction and Ca2+ signal cascades in the apparent Ca2+ requirement of orexin-A signalling. Experimental ApproachReceptor binding was assessed in CHO cells expressing human OX1 receptors with [I-125]-orexin-A by conventional ligand binding as well as scintillation proximity assays. PLC activity was determined by chromatography. Key ResultsBoth orexin receptor binding and PLC activation were strongly dependent on the extracellular Ca2+ concentration. The relationship between Ca2+ concentration and receptor binding was the same as that for PLC activation. However, when Ca2+ entry was reduced by depolarizing the cells or by inhibiting the receptor-operated Ca2+ channels, orexin-A-stimulated PLC activity was much more strongly inhibited than orexin-A binding. Conclusions and ImplicationsCa(2+) plays a dual role in orexin signalling by being a prerequisite for both ligand-receptor interaction and amplifying orexin signals via Ca2+ influx. Some previous results obtained utilizing Ca2+ chelators have to be re-evaluated based on the results of the current study. From a drug discovery perspective, further experiments need to identify the target for Ca2+ in orexin-A-OX1 receptor interaction and its mechanism of action.
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