Cytosolic calcium ion activity in epithelial cells of Necturus kidney

Lee, Chin O.; Taylor, Ann; Windhager, Erich E.

doi:10.1038/287859a0

Cited by 106 publications

(25 citation statements)

References 15 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In the cardiac sarcolemma, this exchange mechanism performs the function of a high capacity system with rapid transport of entered calcium. However, the apparent Km of 8 gM for Na+ is too high for an effective efflux mechanism in a cell whose cytosolic calcium is probably 100-300 nM (23). Using similar techniques to those described in this paper, the original Km for the Na+/Ca2+ exchanger in cardiac sarcolemma vesicles was 8 zM (28), but Caroni et al (30) have shown that with charge equilibration the Km was in fact 1.5 ,uM.…”

Section: Resultsmentioning

confidence: 84%

See 1 more Smart Citation

Calcium transport in canine renal basolateral membrane vesicles. Effects of parathyroid hormone.

Scoble¹,

Mills²,

Hruska³

1985

J. Clin. Invest.

View full text Add to dashboard Cite

IntroductionThe effects of parathyroid hormone were studied on Ca" fluxes in canine renal proximal tubular basolateral membrane vesicles (BLMV). Efflux of Ca2+ from preloaded BLMV was found to be stimulated by an external Na' gradient, and this was inhibited by the Na' ionophore, monensin, and enhanced by intravesicular negative electrical potentials, which indicated electrogenic Na+/Ca2" exchange activity. There was a Na' gradient independent Ca2+ flux, but membrane binding of Ca2" was excluded from contributing to the Na' gradient-dependent efflux. The Na' gradient-dependent flux of Ca2" was very rapid, and even 2-and 5-s points may not fully represent absolute initial rates. It was saturable with respect to the interaction of Ca2" and Na' with an apparent (5 s) K. for Na'-dependent Ca2" uptake of 10 ,uM, and an apparent (5 s) V.,.,, of 0.33 nmol/mg protein per 5 s. The Na' concentration that yielded half maximal Ca2" efflux (2 s) was 11 mM, and the Hill coefficient was two or greater. Both Na+ gradient dependent and independent Ca2" efflux were decreased in BLMV prepared from kidneys of thyroparathyroidectomized (TPTX) dogs, and both were stimulated by parathyroid hormone (PTH) infusion to TPTX dogs. BLMV from TPTX dogs exhibited significantly reduced maximal stimulation of Na+ gradient-dependent Ca2+ uptake with an apparent (5 s) V,,, of 0.23 nmol/mg protein per 5 s, but the apparent K. was 8 ,uM, which was unchanged from normal. The Na+ gradient independent Ca2+ uptake was also reduced in BLMV from TPTX dogs compared with normal. Thus, PTH stimulated both Na+/Ca2" exchange activity and Na+ independent Ca2" flux. In vivo, the latter could result in an elevation of cytosolic Ca2" by PTH, and this might contribute to the observed decrease in solute transport in the proximal tubule. (33). Renal medullary tissue was removed from the kidneys, and renal cortical tissue was diced and homogenized using a Potter-Elvehjem Teflon homogenizer in homogenizing buffer (250 mM sucrose, 20 mM Tris base, and 1 mM EGTA adjusted to pH 7.6), 20 ml/g of tissue.The suspension was then further homogenized using a Polytron homogenizer (Brinkman Instruments, Westbury, NY), setting six for three bursts of 30 s. After samples were taken for analysis of protein and enzyme content, the homogenate was centrifuged at 2,500 g for 15 min and the pellet discarded. The suspension was then spun at 24,000 g for 20 min and the supernatant decanted. The soft portion of the pellet was resuspended in the homogenizing buffer to a value of 1.3 ml/g initial tissue and hand homogenized for seven strokes with a Dounce homogenizer. The suspension was then diluted and thoroughly mixed with 1.6 ml/g initial tissue of 100% Percoll (Pharmacia Fine Chemicals, Upsala, Sweden) and the homogenizing buffer, 17 ml/g initial tissue (final Percoll concentration 8%). The suspension was then centrifuged at 30,000 g for 35 min. This produced a density gradient of Percoll from 1.005 to 1.125 g/ml. The fraction corresponding to an opaque band and density 1.030-1...

show abstract

Section: Resultsmentioning

confidence: 84%

“…In the hamster, Harris et al (13) nephron is unclear. In some respects, the effect of PTH on proximal tubular solute reabsorption is similar to procedures which decrease peritubular Na+ concentrations or are presumed to increase cytosolic Ca2" (23).…”

mentioning

confidence: 75%

Calcium transport in canine renal basolateral membrane vesicles. Effects of parathyroid hormone.

Scoble¹,

Mills²,

Hruska³

1985

J. Clin. Invest.

View full text Add to dashboard Cite

show abstract

“…This discrepancy, which may reflect the decreased buffering with fura-2, has also been observed by others (22,28). However, basal Ca?+ levels measured with both indicators are well within the range of values reported by others using a variety of techniques in epithelial cells (28)(29)(30) as well as other tissues (19, 21, 22, 3 1 Na+-Caz+ exchange has been described in many cell types (32), including epithelial cells (33), and it appears to be involved in regulating basal Ca?+ levels. We tested for the presence of a Na+-Ca2+ exchange process in fura-2-loaded nasal epithelial cells by imposing ion gradients that would be expected to favor N a t for Ca2+,,t exchange.…”

Section: Discussionmentioning

confidence: 80%

Cell Calcium Levels of Normal and Cystic Fibrosis Nasal Epithelium

et al. 1988

View full text Add to dashboard Cite

ABSTRACT. To determine whether epithelial ion transport abnormalities in cystic fibrosis (CF) might reflect abnormal regulation of intracellular Caz+ levels, cytosolic free calcium (Caiz+) was measured using fura-2 or quin2 in suspensions of normal or CF nasal epithelial cells derived from primary cell culture. The basal Ca?+ level measured with fura-2 in CF nasal epithelia was 155 f 9 nM (n = 5), a value not significantly different from normal nasal epithelia (143 f 16 nM, n = 5). Total cell calcium was measured by atomic absorption spectroscopy and no differences were observed between CF (6.3 f 0.5 nmollmg protein; n = 3) and normal (6.2 f 1.2 nmollmg protein; n = 3) nasal epithelial cells. Placing Na+ loaded cells in a low (10 mM) extracellular Na+ solution resulted in a rapid increase in Ca2+ consistent with CaZ+ uptake via a plasmalemmal Na+-Caz+ exchanger. The level of Cai2' achieved by this low Na+ maneuver was not significantly different in CF cells compared to normal cells. Neither isoproterenol (lom5 M) nor forskolin M) had any effect on Ca?+ in normal or CF nasal epithelial cells. Thus, it appears that differences in cell Caiz+, as measured by fluorescent chelators in suspensions of cultured cells, do not account for the abnormalities in basal or isoproterenol stimulated ion transport in CF tissues. (Pediatr Res 24: 79-84,1988) Abbreviations Cai2+, cytolsolic free calcium CF, cystic fibrosis AM, acetoxymethyl ester TCA, trichloroacetic acid PMN, polymorphonuclear cellsThe pathophysiology of CF appears to reflect abnormal epithelial ion transport functions in specific target epithelia (1, 2). Recent evidence has focused on the notion that the abnormalities in epithelial ion transport in the lung reflect dysfunctions in regulatory processes rather than abnormalities in the permeability properties of channel proteins. Evidence for this notion has accrued from two lines of investigation. First, studies of freshly excised nasal epithelia have shown that there is 1) an imbalance between the basal rate of Na+ absorption and the capacity to secrete C1-in CF as compared to normal tissues (3); and 2) the response to P-adrenergic agonists qualitatively differs in the two preparations, i.e., isoproterenol induces C1-secretion in normal

show abstract

“…If, as in other tissues (Blaustein, 1974), this mechanism is rheogenic involving the counterflow of more than 2 Na per Ca, the driving force for Ca extrusion is derived from both the Nachemical gradient and the electrical potential difference across that membrane. There is reasonable evidence that removal of Na from the serosal solution results in an increase in cell Ca and that this may be directly responsible for the decrease in P~'~ observed under these conditions (Grinstein & Erlii, 1978;Lee et al, 1980;Taylor, 1981;Arruda et al, 1982). Thus, it is argued that an increase in (Na)c, which results in a decrease in the electrochemical potential difference for Na across the basolateral membrane, could lead to an increase in cell Ca and that this may be the direct mediator of the "negative feedback" on P~a.…”

Section: Sodium Entry Across the Apical Membranementioning

confidence: 97%

“…Evidence has been presented for the presence of a Na-Ca countertransport mechanism at the basolateral membranes of toad (Chase & A1-Awqati, 1983 ;Arruda et al, 1982) and turtle urinary bladder (Arruda, Sabatini & Westenfelder, 1982), rat small intestine (Hildmann, Schmidt & Murer, 1982), Necturus renal proximal tubule (Lee, Taylor & Windhager, 1980), and frog skin (Grinstein & Erlij, 1978) whereby the movement of Na into the cell across the basolateral membrane energizes the uphill extrusion of Ca from the cell across that barrier. If, as in other tissues (Blaustein, 1974), this mechanism is rheogenic involving the counterflow of more than 2 Na per Ca, the driving force for Ca extrusion is derived from both the Nachemical gradient and the electrical potential difference across that membrane.…”

Section: Sodium Entry Across the Apical Membranementioning

confidence: 98%

Relation between intracellular sodium and active sodium transport in rabbit colon: Current-voltage relations of the apical sodium entry mechanism in the presence of varying luminal sodium concentrations

Turnheim¹,

Thompson²,

Schultz³

1983

J. Membrain Biol.

View full text Add to dashboard Cite

The current-voltage relations of the amiloride-sensitive Na entry pathway across the apical membrane of rabbit descending colon, exposed to a high K serosal solution, were determined in the presence of varying mucosal Na activities, (Na)m, ranging from 6.2 to 99.4 mM. These relations could be closely fit to the "constant field" flux equation yielding estimates of the permeability of the apical membrane to Na, PmNa, and the intracellular Na activity, (Na)c. The following empirical relations emerged: (Na)c increased hyperbolically with increasing (Na)m; PmNa decreased hyperbolically with increasing (Na)m and linearly with increasing (Na)c; spontaneous variations in Na entry rate at constant (Na)m could be attributed entirely to parallel, spontaneous variations in PmNa; the rate of Na entry increased hyperbolically with increasing (Na)m obeying simple Michaelis-Menten kinetics; the relation between (Na)c and "pump rate," however, was sharply sigmoidal and could be fit by the Hill equation assuming strong cooperative interactions between Na and multiple sites on the pump; the Hill coefficient was 2-3 and the value of (Na)c at which the pump-rate is half-maximal was 24 mM. The results provide an internally consistent set of relations among Na entry across the apical membrane, the intracellular Na activity and basolateral pump rate that is also consistent with data previously reported for this and other Na-absorbing epithelia.

show abstract

Cytosolic calcium ion activity in epithelial cells of Necturus kidney

Cited by 106 publications

References 15 publications

Calcium transport in canine renal basolateral membrane vesicles. Effects of parathyroid hormone.

Calcium transport in canine renal basolateral membrane vesicles. Effects of parathyroid hormone.

Cell Calcium Levels of Normal and Cystic Fibrosis Nasal Epithelium

Relation between intracellular sodium and active sodium transport in rabbit colon: Current-voltage relations of the apical sodium entry mechanism in the presence of varying luminal sodium concentrations

Contact Info

Product

Resources

About