Proteins of the TRP family form nonselective Ca²⁺-permeable cation channels. Their activation mechanisms are not completely understood. In particular, the functional properties of TRPM channels, i. e. the members of the melastatin-like TRP subfamily, are still under investigation. Lee et al. (2003) reported a regulation of TRPM3 activity by intracellular calcium stores. These authors cloned cDNAs from human kidney and described six splice variants between 1544 and 1579 amino acids, all starting at the same position as published melastatin does. Recently, we showed the activation of heterologously expressed human TRPM3 by reduction of extracellular osmolarity (Grimm et al., 2003). We cloned the cDNAs of human and mouse TRPM3 from kidney and brain, respectively. The orthologous cDNAs code for a TRPM3 variant with an N-terminal extension of 153 amino acids compared to melastatin and a shorter C-terminus, resulting in a protein of 1325 amino acids. The presence of a very characteristic N-terminal protein motif, found selectively in the sequences of TRPM-channels, makes quite sure that we study a protein identical to the natively expressed TRPM3. By performing 3-prime RACE reactions, we cloned further variants of the TRPM3 protein. The variant with an extended C-terminal end and a length of 1707 amino acids was chosen for expression experiments. In transiently transfected HEK293 cells, we studied the activation mechanism of both TRPM3 variants. We tested for the known stimuli, i.e. spontaneous activity, store-dependent regulation, activation by hypoosmolarity and other extracellular stimuli. In these experiments, the long TRPM3 variant was expressed as determined by Western blot and fluorescence analysis, but did not respond to any of the stimuli used. The short TRPM3 variant (1325 aa) showed spontaneous and hypoosmolarity-induced activity. An activation by intracellular calcium stores, however, was not detectable. These data confirm earlier results of an involvement of TRPM3 in renal osmo-regulation.