A confocal study of mechanism of 5-hydroxytryptamino induced intracellular calcium dynamics in cultured rat stomach fundus smooth muscle cells with a new Ca2+ indicator STDln-AM

A new fluorescent Ca2+ indicator STDln-AM for detecting [Ca2+], transients in cultured smooth muscle cells is presented. By making a comparison, the difference between STDln and fluo-3 is discussed in detail. Using the new Ca2+ indicator, the mechanism of 5-hydroxytryptamino (5-HT) induced intracellular calcium dynamics in stomach fundus smooth muscle cells (SFSMC) of rats is investigated. It is shown that in contrast with fluo-3, STDln is uniformly distributed in the cytosolic compartment but excluded from the nucleus, when it is transfected into cells. This feature makes it a real cytosol Ca2+ indicator and can reflect changes of cytosol [Ca2+] more accurately than that of fluo-3. In addition, STDln responds to the [Ca2+], transients more sensitive and faster than fluo-3. The results also show that, the L-type Ca2+ channel inhibitor Mn9202 and the PLC inhibitor Compound 48/80 can significantly inhibit the [Ca2+], elevation induced by 5-HT, while the PKC inhibitor D-Sphingosine can enhance the effect of 5-HT. The results suggest that 5-HT acts by the way of 5-HT2 receptors on SFSMC, then through 5-HT2 receptors coupled IP3/Ca2+ and GC/PKC double signal transduction pathways to make Ca2+ release from intracellular Ca2+ stores and Ca2+ influx possibly through L-type calcium channels.