Sustained signalling by PTH modulates IP(3) accumulation and IP(3) receptors through cyclic AMP junctions

Parathyroid hormone (PTH) stimulates adenylyl cyclase through type 1 PTH receptors (PTH(1)R) and potentiates the Ca(2+) signals evoked by carbachol, which stimulates formation of inositol 1,4,5-trisphosphate (IP(3)). We confirmed that in HEK cells expressing PTH(1)R, acute stimulation with PTH(1-34) potentiated carbachol-evoked Ca(2+) release. This was mediated by locally delivered cyclic AMP (cAMP), but unaffected by inhibition of protein kinase A (PKA), exchange proteins activated by cAMP, cAMP phosphodiesterases (PDEs) or substantial inhibition of adenylyl cyclase. Sustained stimulation with PTH(1-34) causes internalization of PTH(1)R–adenylyl cyclase signalling complexes, but the consequences for delivery of cAMP to IP(3)R within cAMP signalling junctions are unknown. Here, we show that sustained stimulation with PTH(1-34) or with PTH analogues that do not evoke receptor internalization reduced the potentiated Ca(2+) signals and attenuated carbachol-evoked increases in cytosolic IP(3). Similar results were obtained after sustained stimulation with NKH477 to directly activate adenylyl cyclase, or with the membrane-permeant analogue of cAMP, 8-Br-cAMP. These responses were independent of PKA and unaffected by substantial inhibition of adenylyl cyclase. During prolonged stimulation with PTH(1-34), hyperactive cAMP signalling junctions, within which cAMP is delivered directly and at saturating concentrations to its targets, mediate sensitization of IP(3)R and a more slowly developing inhibition of IP(3) accumulation.