Modulation of Na+/Ca2+ exchange in epithelial cells of porcine thick ascending limb

greedy — Sat, 12 Sep 2009 00:57:00 +0000

Dai LJ, Ritchie G, Bapty B, Auger V, and Quamme GA: Modulation of Na⁺/Ca²⁺exchange in epithelial cells of porcine thick ascending limb. American Journal of Physiology 270:F953-F959, 1996.

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Abstract Modulation of Nat/ Ca2+ exchange in epithelial cells of porcine thick ascending limb. Am. J. Physiol. 270 (Renal Fluid Electrolyte Physiol. 39): F953-F959, 1996.-We have provided functional and molecular evidence for the presence of Na+/Ca2+ exchange in isolated porcine cortical thick ascending limb (CTAL) cells. The present studies were designed to show that this exchange activity may be modulated by phosphorylative processes. Control of intracellular Ca2+ concentration ( [Ca2+];) was determined in isolated CTAL cells with microfluorescence. CTAL cells were pretreated with ouabain to elevate intracellular Na+ concentration ([Na+]i) from 10 to 20 mM. These cells had normal basal [Ca2+]; (79 2 3 nM). Substitution of extracellular NaCl (50 mM) with KC1 resulted in the rapid elevation of [Ca2+]; to maximal levels of 795 t 60 nM (n = 17). The increments of [Ca2+]; were associated with [Na+]i. We next determined the modulation of Na+lCa2+ exchange activity with phosphorylative inhibitors. Pretreatment of cells with calmidazolium, a Ca2+- calmodulin inhibitor, resulted in a shift of the [Na+]i dependence curve to the right. Pretreatment with okadaic acid, a phosphatase 1 and 2A inhibitor, increased the Na+/Ca2+ exchanger activity resulting in halfmaximal [Ca2+]; increase near normal [Na+]i of 12 mM. Furthermore, in the presence of okadaic acid in normal CTAL cells, pretreatment with ouabain and the elevation of [Na+]i was not required to elicit increments in [Ca2+];. These data indicate that Na+/Ca2+ exchange is present in CTAL cells and the exchange activity appears to be modulated, directly or indirectly, by phosphorylation events.

Modulation of Nat/

Ca2+ exchange in epithelial cells of porcine thick ascending

limb. Am. J. Physiol. 270 (Renal Fluid Electrolyte Physiol.

39): F953-F959, 1996.-We have provided functional and

molecular evidence for the presence of Na+/Ca2+ exchange in

isolated porcine cortical thick ascending limb (CTAL) cells.

The present studies were designed to show that this exchange

activity may be modulated by phosphorylative processes.

Control of intracellular Ca2+ concentration ( [Ca2+];) was

determined in isolated CTAL cells with microfluorescence.

CTAL cells were pretreated with ouabain to elevate intracellular

Na+ concentration ([Na+]i) from 10 to 20 mM. These

cells had normal basal [Ca2+]; (79 2 3 nM). Substitution of

extracellular NaCl (50 mM) with KC1 resulted in the rapid

elevation of [Ca2+]; to maximal levels of 795 t 60 nM (n = 17).

The increments of [Ca2+]; were associated with [Na+]i. We

next determined the modulation of Na+lCa2+ exchange activity

with phosphorylative inhibitors. Pretreatment of cells

with calmidazolium, a Ca2+- calmodulin inhibitor, resulted in

a shift of the [Na+]i dependence curve to the right. Pretreatment

with okadaic acid, a phosphatase 1 and 2A inhibitor,

increased the Na+/Ca2+ exchanger activity resulting in halfmaximal

[Ca2+]; increase near normal [Na+]i of 12 mM.

Furthermore, in the presence of okadaic acid in normal CTAL

cells, pretreatment with ouabain and the elevation of [Na+]i

was not required to elicit increments in [Ca2+];. These data

indicate that Na+/Ca2+ exchange is present in CTAL cells and

the exchange activity appears to be modulated, directly or

indirectly, by phosphorylation events.

intracellular calcium ion concentration; intracellular sodium

Atrial natriuretic peptide initiates Ca2+ transients in isolated renal cortical thick ascending limb cells

greedy — Sat, 12 Sep 2009 00:50:18 +0000

Dai LJ, and Quamme GA: Atrial natriuretic peptide initiates Ca²⁺ transients in isolated renal cortical thick ascending limb cells. American Journal of Physiology 265:F592-F597, 1993.

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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.

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.

porcine thick ascendingAtria1 natriuretic