Fig. 1.

Regulation of IP₃Rs by cAMP signalling junctions.

(A) Populations of fluo4-loaded HEK-293 cells stably expressing type 1 receptors for PTH were stimulated with PTH (100 nM, red line) and carbachol (20 μM, both lines) as indicated after addition of BAPTA to chelate extracellular Ca²⁺. Results show that PTH does not alone evoke an increase in [Ca²⁺]_c, but it potentiates the Ca²⁺ signal evoked by carbachol. Similar results are shown in Refs. [36], [39]. (B) In HEK-293 cells, PTH through heterologously expressed type 1 PTH receptors stimulates adenylyl cyclase and so formation of cAMP. Carbachol stimulates endogenous M₃ muscarinic receptors, which activate PLC and thereby formation of IP₃ and release of Ca²⁺ from the ER through IP₃Rs. The potentiation of carbachol-evoked Ca²⁺ signals by PTH is entirely mediated by cAMP, which enhances IP₃-evoked Ca²⁺ release from IP₃Rs. (C) IP₃R2 and AC6 are selectively associated at cAMP signalling junctions. Within these junctions, cAMP is delivered from AC to IP₃Rs at concentrations far greater than needed to maximally sensitize the associated IP₃Rs. This allows each junction to function as a digital ‘on-off’ switch, it ensures that cAMP-mediated signalling operates with a considerable safety margin, and it allows rapid activation of the associated IP₃Rs (as cAMP is locally delivered at high concentration) and rapid de-activation (as diffusion of cAMP form the junction reduces its concentration to below that needed for sensitizing IP₃Rs). Since each signalling junction operates as an ‘on-off’ switch, the concentration-dependent effects of PTH are proposed to be due to recruitment of junctions, rather than to graded activity within individual junctions.