Expression and function of ryanodine receptors in human melanocytes

We investigated the expression of ryanodine receptors (RyRs) in cultured hu man melanocytes with immunocytochemistry and reverse transcriptase-polymera se chain reaction. With the use of a monoclonal antibody, RyR immunoreactiv ity was detected in the cytoplasm of melanocytes, and was further confirmed by RT-PCR assay. The PCR products were cut with restriction enzymes specif ic for each RyR isoform. Using the RyR1-specific restriction enzyme Sad yie lded fragments of 300. 100. and 130 base pairs, consistent with the express ion of RyR1 irs, consistent with the expression of RyR1 isoforms. The funct ion of RyR in Ca2+ signaling was investigated using single-cell fura-2 imag ing. Ryanodine (1 to similar to 100 mu M) induced significant elevation of cytoplasmic Ca2+ in single human melanocytes in a dose;dependent manner. Th e ryanodine-induced [Ca2+](i) increase was inhibited by neomycin. Furthermo re, ryanodine inhibited proliferation and stimulated pigmentation of human melanocytes. This study demonstrates that the RyR1 isoform is expressed in cultured human melanocytes, and suggests that the RyR may be involved in re gulating the intracellular Ca2+ responses involved in proliferation and pig mentation of cultured human melanocytes. (C) 2000 Wiley-Liss, Inc.