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Na+/Ca2+ exchanger in epithelial cells of the porcine cortical thick ascending limb. Am. J. Physiol. 270 (Renal Fluid Electrolyte Physiol. 39): F411-F418,1996.-Intracellular Ca2+ concentration ( [Ca2+]i) plays an important role in the signal transduction processes within cortical thick ascending limb (CTAL) cells. Control of [Ca2+]; was investigated in isolated CTAL cells with microfluorescent techniques. CTAL cells pretreated with ouabain to elevate intracellular Na+ concentration ([Na+]i) had basal [Ca2+]. 10 f 86 IT 2 nM. Removal of extracellular Na (Na,f ) or voltage depolarization with KC1 (in the presence of Na,f) resulted in a rapid and reversible maximal elevation of [Ca2+]i (1,023 t 72 nM, n = ZS), which was dependent on the presence of external Ca 2+ (Caf’). The rise in [Ca2+]i was inhibited with La3+, Mg2+, amiloride, and bepridil. The increments of [Ca2+]i with either removal of Naz or voltage depolarization were dependent on pretreatment with ouabain and increases in [Na+]i. The presence of a Na+/Ca2+ exchanger was confirmed with hybridization techniques, and the isoform was identified by sequencing the alternative splicing site within the intracellular loop. A gene transcript that encodes a portion of the intracellular loop of the renal Na+/Ca2+ exchanger was amplified from cortical tissue and single CTAL cells by reverse transcription-polymerase chain reaction, using primers flanking the alternative splicing site. Southern hybridization and DNA sequencing demonstrated the isoform contained exons B and D, which is characteristic of one isoform (NACA3) of the renal Na+/Ca2+ exchanger. The results provide both functional and molecular evidence for a Na+/Ca2+ exchanger in thick ascending limb cells of the porcine kidney.