We have recently demonstrated that the cells expressing CD36, localized apically on the taste buds of mouse lingual circumvallate papillae (CVP), act as gustatory cells. In the present study, we isolated these CD36-positive cells from mouse CVP and investigated intracellular signaling events, triggered by a long-chain polyunsaturated fatty acid, i.e., linoleic acid (LA). LA induced increases in free intracellular calcium concentrations, [Ca2+]i, by recruiting calcium from endoplasmic reticulum (ER) pool via IP3 production, followed by calcium influx via opening of store operated calcium (SOC) channels. LA also induced phosphorylation of Src-protein tyrosine kinases (Src-PTKs), particularly of Fyn59 and Yes62. LA-evoked phosphorylation of Fyn59 and Yes62 was implicated in the activation of SOC channels. RT-qPCR revealed that the CD36-positive gustatory cells possessed mRNA of enzymes like tryptophan hydroxylase-1 (TPH-1), L-aromatic amino acid decarboxylase (AADC), tyrosine hydroxylase (TH), dopamine beta-hydroxylase (DBH), involved in the synthesis of monoamine neurotransmitters. Interestingly, addition of LA to these cells induced the release of 5-HT and noradrenalin (NA) to the extracellular environment. The LA-induced release of these neurotransmitters was curtailed by SOC channel blockers and Src-PTK inhibitors. These results altogether demonstrate that LA binds to mouse CD36-positive gustatory cells, induces Src-PTKs phosphorylation, triggers calcium signaling and evokes the release of 5-HT and NA which, in turn, may be implicated in the downstream signaling to the afferent nerve fibers, thus transmitting the output signal from taste buds to the central nervous system.