Real time computerized intracellular calcium ion imaging: a rapid method of neuronal receptor evaluation

Real time computerized intracellular calcium ion analysis using fura-2-based fluorescence imaging has been used to evaluate neurotransmitter receptor function in primary cultures of rat hypothalamic neurons and a rat hypothalamic neuronal cell line (LT6/4). A wide range of neurotransmitters induced an increase in intracellular Ca2+ in cultured hypothalamic neurons. In contrast, undifferentiated LT6/4 cells failed to response to 56mM K+ or to a battery of neurotransmitters with the exception of histamine (100microM). LT6/4 cells, induced to differentiate by prior exposure to dibutyryl cAMP and NGF, responded to K+ but lost their responsiveness to histamine. Computerized imaging allowed the intracellular Ca2+ response of up to 50 cells to be determined simultaneously without resort to complex electrophysiological manipulations or receptor binding studies. The assessment of receptor complement and coupling using this technique allows large amounts of reproducible data to be rapidly collected from small populations of cells.