We wished to document whether the SV40 immortalized rat submanbidular cell line had preserved their intracellular Ca²⁺ response to adequate neurotransmitters and determine if it would lend itself to further comparative cell physiological studies. The cell line, a gift from D Quissell, U of Colorado, was grown under standard cell culture conditions. At the time of experiments, cells were incubated with fura-2-AM (Molecular Probes, USA) following established routines. The cells were studied in a flow through chamber with an inverted microscope. As a measure of the intracellular Ca²⁺ concentration, fura-2 was excited at 510 nm and the ratio of emitted light at 340 and 380 nm was recorded using MetaFluor 7.1.2 software (Universal Imaging, USA). After a few minutes in an isosmotic solution (300 mOsm, pH 7.4), the cells were exposed to norepinephrine 10^­5M, carbamoylcholine chloride 10^­5M, ATP 10^­4 M and substance P 10^­7 M for 15 min and the changes in intracellular Ca²⁺ were measured. Exposure to all neurotransmitters induced a transient increase in the intracellular Ca²⁺-level (norepinephrine 185±18% carbamoylcholine chloride 426±1%, substance P 230±38%, and ATP 127±27%. The peak Ca²⁺-increase occurred at 11 sec (norepinephrine and carbamoylcholine chloride), 90 sec (substance P) and 18 sec (ATP) following transmitter exposure. While norepinephrine and carbamoylcholine chloride increased the Ca²⁺-concentration in all cells measured, substance P and ATP induced Ca²⁺- responses in about 50% of the measured cells. In conclusion, in this cell line there was a strong intracellular Ca²⁺-increase in response to norepinephrine and carbamoylcholine chloride and a variable response to ATP and substance P. Thus, this cell line has retained a normal Ca²⁺-response to transmitters of the autonomic nervous system.