Inositol 1,4,5-trisphosphate and calcium regulate the calcium channel function of the hepatic inositol 1,4,5-trisphosphate receptor

The regulation of the inositol 1,4,5-trisphosphate (IP3) receptor in liver was analyzed using a novel superfusion method. Hepatic microsomes were loaded with Ca-45(2+), and superfused at high flow rates to provide precise control over IP3 and Ca2+ concentrations ([Ca2+]) and to isolate Ca-45(2+) release from reuptake. Ca-45(2+) release was dependent on both [Ca2+] and IP3. The initial rate of Ca-45(2+) release was a biphasic function of [Ca2+], increasing as [Ca2+] approached 3 mu M but decreasing at higher concentrations, suggesting that the hepatic IP3 receptor is regulated by [Ca2+] at two sites, a high affinity potentiation site and a low affinity inhibitory site. The relationship between initial rates and IP3 concentration was steep (Hill coefficient of 3.4), suggesting that activation of the calcium channel requires binding of at least 3 IP3 molecules. IP3 concentrations above 10 mu M produced rapid decay of release rates, suggesting receptor inactivation. Superfusion with 10 mu M IP3 under conditions that minimize calcium release ([Ca2+] < 1 mM) inhibited Ca-45(2+) release in response to subsequent stimulation (400 nM Ca2+). These data suggest sequential positive and negative regulation of the hepatic IP3 receptor by cytosolic calcium and by IP3, which may underlie hepatocellular propagation of regenerative, oscillatory calcium signals.