Activation of Na/H exchanger in mesangial cells is associated with translocation of PKC isoforms

Ramesh Saxena, Brett A. Saksa, Alan P. Fields, Michael B. Ganz

Research output: Contribution to journalArticle

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Abstract

We investigated the potential coupling of the Na/H ex changer to protein kinase C (PKC) activation. Mesangial cells (MC), passage 3-8, were exposed to either serotonin (5-HT), arginine vasopressin (AVP), or fibroblast growth factor (FGF). We assessed the effect of these agonists on recovery from an acid load (NH4 +/NH3) in the nominal absence of CO2/HCO3. 5-HT, AVP, and FGF significantly enhanced the rate of recovery from 24.2 ±4.5 × 10-4 intracellular pH units/s to 67.7 ± 5.7, 70.5 ± 5.1, and 55.7 ± 6.8, respectively (n >6, P< 0.0001). The increase in activity was abolished by ethylisopropylamiloride and removal of extracellular Na+, thereby suggesting that activation of Na/H exchanger activity was responsible for enhanced recovery by 5-HT, AVP, and FGF. MC were then pretreated with phorbol ester [phorbol 12-myristate 13-acetate (PMA); 10 μM for 40 h] and acid loaded. 5-HT and AVP did not enhance recovery of PMA-pretreated cells (23.3 ±6.8 and 24.9 ± 4.7, n > 4, not significant), whereas FGF stimulated activity (48.2 ± 5.5, n > 4, P < 0.001). Similar results were found when MC were pretreated with the PKC antagonist, sphingosine. Specific antibodies were raised against α-, βI-, βII-, and γ-isoforms of PKC. We observed that MC express the α-, γ-, and βIisoforms, but not the βII-isoform of PKC. When MC were exposed to 5-HT and AVP, translocation was evident immediately (within seconds) for the α-isoform, whereas maximal translocation occurred at 30 min for βIPKC and at 24 h for -γ-PKC (n > 6 for each agonist). FGF did not induce translocation of any of the PKC isoforms studied. These results indicate that in MC, PKC activation is required for the 5-HT and AVP stimulation of the Na/H exchanger. There appears to be a temporal correlation between the translocation pattern of the α-isoform of PKC and Na/H exchange activation.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Renal Fluid and Electrolyte Physiology
Volume265
Issue number1 34-1
StatePublished - 1993

Fingerprint

Sodium-Hydrogen Antiporter
Mesangial Cells
Fibroblast Growth Factors
Protein Kinase C
Arginine Vasopressin
Serotonin
Protein Isoforms
Acids

Keywords

  • Arginine vasopressin
  • Fibroblast growth factor
  • Protein kinase c isoforms
  • Serotonin
  • Sodium-hydrogen exchanger

ASJC Scopus subject areas

  • Physiology

Cite this

Activation of Na/H exchanger in mesangial cells is associated with translocation of PKC isoforms. / Saxena, Ramesh; Saksa, Brett A.; Fields, Alan P.; Ganz, Michael B.

In: American Journal of Physiology - Renal Fluid and Electrolyte Physiology, Vol. 265, No. 1 34-1, 1993.

Research output: Contribution to journalArticle

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abstract = "We investigated the potential coupling of the Na/H ex changer to protein kinase C (PKC) activation. Mesangial cells (MC), passage 3-8, were exposed to either serotonin (5-HT), arginine vasopressin (AVP), or fibroblast growth factor (FGF). We assessed the effect of these agonists on recovery from an acid load (NH4 +/NH3) in the nominal absence of CO2/HCO3. 5-HT, AVP, and FGF significantly enhanced the rate of recovery from 24.2 ±4.5 × 10-4 intracellular pH units/s to 67.7 ± 5.7, 70.5 ± 5.1, and 55.7 ± 6.8, respectively (n >6, P< 0.0001). The increase in activity was abolished by ethylisopropylamiloride and removal of extracellular Na+, thereby suggesting that activation of Na/H exchanger activity was responsible for enhanced recovery by 5-HT, AVP, and FGF. MC were then pretreated with phorbol ester [phorbol 12-myristate 13-acetate (PMA); 10 μM for 40 h] and acid loaded. 5-HT and AVP did not enhance recovery of PMA-pretreated cells (23.3 ±6.8 and 24.9 ± 4.7, n > 4, not significant), whereas FGF stimulated activity (48.2 ± 5.5, n > 4, P < 0.001). Similar results were found when MC were pretreated with the PKC antagonist, sphingosine. Specific antibodies were raised against α-, βI-, βII-, and γ-isoforms of PKC. We observed that MC express the α-, γ-, and βIisoforms, but not the βII-isoform of PKC. When MC were exposed to 5-HT and AVP, translocation was evident immediately (within seconds) for the α-isoform, whereas maximal translocation occurred at 30 min for βIPKC and at 24 h for -γ-PKC (n > 6 for each agonist). FGF did not induce translocation of any of the PKC isoforms studied. These results indicate that in MC, PKC activation is required for the 5-HT and AVP stimulation of the Na/H exchanger. There appears to be a temporal correlation between the translocation pattern of the α-isoform of PKC and Na/H exchange activation.",
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T1 - Activation of Na/H exchanger in mesangial cells is associated with translocation of PKC isoforms

AU - Saxena, Ramesh

AU - Saksa, Brett A.

AU - Fields, Alan P.

AU - Ganz, Michael B.

PY - 1993

Y1 - 1993

N2 - We investigated the potential coupling of the Na/H ex changer to protein kinase C (PKC) activation. Mesangial cells (MC), passage 3-8, were exposed to either serotonin (5-HT), arginine vasopressin (AVP), or fibroblast growth factor (FGF). We assessed the effect of these agonists on recovery from an acid load (NH4 +/NH3) in the nominal absence of CO2/HCO3. 5-HT, AVP, and FGF significantly enhanced the rate of recovery from 24.2 ±4.5 × 10-4 intracellular pH units/s to 67.7 ± 5.7, 70.5 ± 5.1, and 55.7 ± 6.8, respectively (n >6, P< 0.0001). The increase in activity was abolished by ethylisopropylamiloride and removal of extracellular Na+, thereby suggesting that activation of Na/H exchanger activity was responsible for enhanced recovery by 5-HT, AVP, and FGF. MC were then pretreated with phorbol ester [phorbol 12-myristate 13-acetate (PMA); 10 μM for 40 h] and acid loaded. 5-HT and AVP did not enhance recovery of PMA-pretreated cells (23.3 ±6.8 and 24.9 ± 4.7, n > 4, not significant), whereas FGF stimulated activity (48.2 ± 5.5, n > 4, P < 0.001). Similar results were found when MC were pretreated with the PKC antagonist, sphingosine. Specific antibodies were raised against α-, βI-, βII-, and γ-isoforms of PKC. We observed that MC express the α-, γ-, and βIisoforms, but not the βII-isoform of PKC. When MC were exposed to 5-HT and AVP, translocation was evident immediately (within seconds) for the α-isoform, whereas maximal translocation occurred at 30 min for βIPKC and at 24 h for -γ-PKC (n > 6 for each agonist). FGF did not induce translocation of any of the PKC isoforms studied. These results indicate that in MC, PKC activation is required for the 5-HT and AVP stimulation of the Na/H exchanger. There appears to be a temporal correlation between the translocation pattern of the α-isoform of PKC and Na/H exchange activation.

AB - We investigated the potential coupling of the Na/H ex changer to protein kinase C (PKC) activation. Mesangial cells (MC), passage 3-8, were exposed to either serotonin (5-HT), arginine vasopressin (AVP), or fibroblast growth factor (FGF). We assessed the effect of these agonists on recovery from an acid load (NH4 +/NH3) in the nominal absence of CO2/HCO3. 5-HT, AVP, and FGF significantly enhanced the rate of recovery from 24.2 ±4.5 × 10-4 intracellular pH units/s to 67.7 ± 5.7, 70.5 ± 5.1, and 55.7 ± 6.8, respectively (n >6, P< 0.0001). The increase in activity was abolished by ethylisopropylamiloride and removal of extracellular Na+, thereby suggesting that activation of Na/H exchanger activity was responsible for enhanced recovery by 5-HT, AVP, and FGF. MC were then pretreated with phorbol ester [phorbol 12-myristate 13-acetate (PMA); 10 μM for 40 h] and acid loaded. 5-HT and AVP did not enhance recovery of PMA-pretreated cells (23.3 ±6.8 and 24.9 ± 4.7, n > 4, not significant), whereas FGF stimulated activity (48.2 ± 5.5, n > 4, P < 0.001). Similar results were found when MC were pretreated with the PKC antagonist, sphingosine. Specific antibodies were raised against α-, βI-, βII-, and γ-isoforms of PKC. We observed that MC express the α-, γ-, and βIisoforms, but not the βII-isoform of PKC. When MC were exposed to 5-HT and AVP, translocation was evident immediately (within seconds) for the α-isoform, whereas maximal translocation occurred at 30 min for βIPKC and at 24 h for -γ-PKC (n > 6 for each agonist). FGF did not induce translocation of any of the PKC isoforms studied. These results indicate that in MC, PKC activation is required for the 5-HT and AVP stimulation of the Na/H exchanger. There appears to be a temporal correlation between the translocation pattern of the α-isoform of PKC and Na/H exchange activation.

KW - Arginine vasopressin

KW - Fibroblast growth factor

KW - Protein kinase c isoforms

KW - Serotonin

KW - Sodium-hydrogen exchanger

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