Ryanodine Receptors in Liver

Abstract

The ryanodine receptor has been mainly regarded as the Ca 2؉ release channel from sarcoplasmic reticulum controlling skeletal and cardiac muscle contraction. However, many studies have shown that it is widely expressed, with functions not restricted to muscular contraction. This study examined whether ryanodine receptor plays a role in calcium signaling in the liver. RT-PCR analysis of isolated hepatocytes showed expression of a truncated type 1 ryanodine receptor, but no type 2 or type 3 message was detected. We also detected binding sites for [3 H]ryanodine in the microsomal cellular fraction and in permeabilized hepatocytes. This binding was displaced by caffeine and dantrolene, but not by ruthenium red, heparin or cyclic ADPRibose. Ryanodine, by itself, did not trigger Ca 2؉ oscillations in either primary cultured hepatocytes or hepatocytes within the intact perfused rat liver. In both preparations, however, ryanodine significantly increased the frequency of the cytosolic free [Ca 2؉ ] oscillations evoked by an ␣ 1 adrenergic receptor agonist. Experiments in permeabilized hepatocytes showed that both ryanodine and cyclic ADP-ribose evoked a slow Ca 2؉ leak from intracellular stores and were able to increase the Ca 2؉ -released response to a subthreshold dose of inositol 1,4,5-trisphosphate. Our findings suggest the presence of a novel truncated form of the type 1 ryanodine receptor in rat hepatocytes. Ryanodine modulates the pattern of cytosolic free [Ca 2؉ ] oscillations by increasing oscillation frequency. We propose that the Ca 2؉ released from ryanodine receptors on the endoplasmic reticulum provides an increased pool of Ca 2؉ for positive feedback on inositol 1,4,5-trisphosphate receptors.Two families of channels lead to Ca 2ϩ release from intracellular stores in the endoplasmic reticulum (ER) 2 /sarcoplasmic reticulum (SR). One is the inositol 1,4,5-trisphosphate receptor (IP 3 R) family. Three different isoforms (types I, II, III) have been characterized and there are also a number of splicing variants. Activation of the IP 3 Rs is elicited by IP 3 generated by receptor activation of phosphoinositide-specific phospholipase C (PLC). The IP 3 Rs are predominantly situated in the ER and play a pivotal role in Ca 2ϩ signaling in almost every cell type (1, 2). The second family of intracellular Ca 2ϩ channels has been named ryanodine receptor (RyR) because of its high affinity for the plant alkaloid ryanodine that led to its identification and purification (3). Ryanodine has been utilized as a tool to study Ca 2ϩ release from the ER/SR of, at first, muscle cells, and more recently of a wide variety of nonmuscle cells. As with the IP 3 R family, the RyR family consists of three isoforms: RyR1 is commonly associated with skeletal muscle cells, RyR2 with cardiac muscle cells and RyR3 is widely distributed with low expression and unclear physiological role (4).An interesting feature of both channel families is their regulation by Ca 2ϩ itself. Ca 2ϩ can either stimulate or inhibit the channels depending on...

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