Two Types of Cellular Serotonin Biosensors

The method of cellular biosensors, which allows local detection of substances secreted by single cells, is traditionally used for studying the peripheral taste system. This method is based on cells that can detect secreted molecules with surface receptors coupled to the mobilization of intracellular Ca²⁺. Thus, the neurotransmitter release can be tracked on-line by Ca²⁺ signals generated by a cell-biosensor. A specific feature of such cellular biosensors is that Са²⁺ signals induced by GPCR ligands may be generated in the “all-or-nothing” manner due to the involvement of trigger-like mechanism, that is Ca²⁺-induced Ca²⁺ release (CICR). Although such a sensor validates the fact of neurotransmitter secretion, it does not allow studying regulatory circuits controlling the quantity of the released substance. This implies that biosensors, which are based on distinct intracellular signaling pathways, such as for cAMP signaling, could be more informative. Here we generated two types of serotonin biosensors. One was based on CHO cells expressing the recombinant 5-HT2C receptor coupled by the phospholipase pathway to Ca²⁺ mobilization. Another involved HEK-293 cells that expressed both 5-HT2C receptor coupled to adenylyl cyclase and the fluorescent protein Pink Flamindo serving as genetically encoded cAMP sensor. Although both cellular biosensors allow detection of nanomolar serotonin, 5-НТ2С cells generated Ca²⁺ responses in the “all-or-nothing” manner. In contrast, in 5-HT4 and Pink Flamindo expressing cells, serotonin-triggered cAMP responses gradually rose with agonist concentration in the range of 0.3–30 nM. It thus appears that the 5-НТ4/Pink Flamindo biosensor is suitable for studying regulatory mechanism of serotonin secretion.