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Abstract Atria1 natriuretic peptide initiates Ca2+ transients in isolated renal cortical thick ascending limb cells. Am. J. Physiol. 265 (Renal Fluid Electrolyte PhysioZ. 34): F592-F597, 1993.-The following studies identified and characterized atria1 natriuretic peptide (ANP) receptor-mediated Ca2+ transients in cortical thick ascending limb (CTAL) cells. Primary cell cultures were prepared from porcine kidneys by immunodissection, and intracellular Ca2+ concentration ([Ca”+]J was determined in single cells with microfluorometry. ANP (10m7M ) and its analogue,C -type natriuretic peptide (CNP, 10m7M ), elicited Ca2+t ransients [ 104 t 6 (basal levels) to 653 & 112 nM (stimulated) and from 84 t 4 to 209 & 18 nM, respectively]. Receptor-mediated [ Ca2+]i increase was dose-dependenwt ith a 50% effective concentration (EC,,) of N lo-lo M. The increment in [Ca”‘]i was due to internal release and influx across the plasma membrane. Prior treatment of ANP or CNP (10q7 M) did not markedly affect a post application of either ANP or CNP. The truncated analogue of ANP, C-ANP-(4-23), which preferentially binds to clearance receptors, elicited an increase in [Ca”+]i (82 & 1 to 427 t 41 nM). 8-Bromoguanosine 3’,5’-cyclic monophosphate (8-BrcGMP) did not alter [Ca”+]i, but pretreatment of CTAL cells with 8-BrcGMP for 30 min before agonist treatment prevented ANP-induced Ca2+ signals [83 t 5 (basal) to 88 t 5 nM (stimulated)]. These results are evidence for the existence of clearance ANP receptors in CTAL cells that may have biological functions and clearance. The functional responseso f these signal interactions may have important consequenceosn hormone actions with the CTAL.

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Abstract Cyclic nucleotides alter intracellular free Mg2+ in renal epithelial cells. Am. J. Physiol. 262 (Renal Fluid Electrolyte Physiol. 31): FllOOF1104,1992.- Intracellular Mg”+ plays an important role in cell physiology. Studies were performed on MDCK cells and primary cortical thick ascending limb (CTAL) cells to determine hormonal influences on intracellular Mg2+ control. Free Mg2+ ( [Mg2+]i) was measured by fluorescence with mag-fura-2. Addition of 8bromoguanosine 3’,5’-cyclic monophosphate @-BrcGMP, 10e4 M) to subconfluent MDCK cells resulted in rapid increases in [Mg2+]i from basal levels of 552 it 6 PM to peak concentrations of 682 t 5 PM, whereas 8BrcAMP (10v4 M) led to significant decreases in [Mg2+]i from 538 of 5 to 362 t 17 PM. These effects of cyclic nucleotides were dose dependent with half-maximal concentrations (EC,,) of – 10e5 M for both increments in [Mg2+]i with cGMP and decrements in [Mg”+]i with CAMP. Atria1 natriuretic peptide (ANP) and cGMP increased Mg2+ in porcine primary CTAL cells from 525 t 12 to 592 & 18 PM and from 538 & 8 to 609 t 18 PM, respectively. The increment in [Mg2+]i with ANP was dose responsive with EC& values of – lo-l1 M suggesting that these effects may be of physiological importance. Parathyroid hormone and calcitonin and their second messenger, CAMP, diminished Mg2+ by -80 PM. The EC& value for calcitonin was in the order of 10mg M. The changes in [Mg2+]i, whether increases with ANP or cGMP and decreases with PTH, calcitonin, or CAMP, were rapid in nature and independent of changes in intracellular free Ca2+ concentration. These data indicate that [Mg”+]i is influenced by peptide hormones and their second messengers likely through activation of appropriate protein kinases. The functional role of these changes remains to be determined.