Abstract
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 45Ca2+, and superfused at high flow rates to provide precise control over IP3 and Ca2+ concentrations ([Ca2+]) and to isolate 45Ca2+ release from reuptake. 45Ca2+ release was dependent on both [Ca2+] and IP3. The initial rate of 45Ca2+ release was a biphasic function of [Ca2+], increasing as [Ca2+] approached 3 microM 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 microM produced rapid decay of release rates, suggesting receptor inactivation. Superfusion with 10 microM IP3 under conditions that minimize calcium release ([Ca2+] < 1 nM) inhibited 45Ca2+ 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.
Highlights
The intracellular receptor for IP3 is a calcium channel that permits efflux of Ca2ϩ sequestered in the endoplasmic reticulum out into the cytoplasm
The aim of this study was to examine the regulation of the IP3 receptor by two second messengers implicated in its function, IP3 and Ca2ϩ
Our results are consistent with a model for the IP3 receptor where either Ca2ϩ or IP3 are sufficient to promote receptor inactivation, but both are necessary to promote receptor activation
Summary
In many cells including hepatocytes, binding of certain hormones to cell surface receptors results in the activation of phospholipase C, thereby generating the intracellular second messenger inositol 1,4,5-trisphosphate (IP3). The intracellular receptor for IP3 is a calcium channel that permits efflux of Ca2ϩ sequestered in the endoplasmic reticulum out into the cytoplasm. Ca2ϩ activates the release process, whereas, at higher concentrations, Ca2ϩ is inhibitory This regulation by calcium has been proposed to be a means of controlling calcium oscillations [1, 2]. A superfusion assay, which was developed to study IP3-dependent calcium release from brain microsomes [5], was adapted for use with liver microsomes This assay achieves subsecond time resolution, allows for precise control of extravesicular Ca2ϩ and IP3 concentrations, and isolates the release of 45Ca2ϩ from its reuptake. We show that both extravesicular Ca2ϩ and IP3 can activate and inhibit the calcium channel function of the IP3 receptor
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
