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K depletion enhances the extracellular Ca2+-induced inhibition of the apical K channels in the mTAL of rat kidney.

Gu RM, Wei Y, Jiang HL, Lin DH, Sterling H, Bloom P, Balazy M, Wang WH - J. Gen. Physiol. (2002)

Bottom Line: Physiol. 270:C103-C111).Also, addition of S-nitroso-N-acetylpenicillamine abolished the inhibitory effect of arachidonic acid on channel activity in the mTAL, whereas it did not block the inhibitory effect of 20-HETE.We conclude that a low dietary K intake increases the sensitivity of the 70-pS K channel to the extracellular Ca(2)+, and that a decrease in NOS activity is involved in enhancing the inhibitory effect of the extracellular Ca(2)+ on channel activity in the mTAL during K depletion.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, New York Medical College, Valhalla, NY 10595, USA.

ABSTRACT
We have shown previously that raising extracellular Ca(2)+ inhibited the apical 70-pS K channel in the thick ascending limb (TAL; Wang, W.H., M. Lu, and S.C. Hebert. 1996. Am. J. Physiol. 270:C103-C111). We now used the patch-clamp technique to study the effect of increasing the extracellular Ca(2)+ on the 70-pS K channel in the mTAL from rats on a different K diet. Increasing the extracellular Ca(2)+ from 10 microM to 0.5, 1, and to 1.5 mM in the mTAL from rats on a K-deficient (KD) diet inhibited the channel activity by 30, 65, and 90%, respectively. In contrast, raising the extracellular Ca(2)+ to 1.5 mM had no significant effect on channel activity in the mTAL from animals on a high K (HK) diet and further increasing the extracellular Ca(2)+ to 2.5, 3.5, and 5.5 mM decreased the channel activity by 29, 55, and 90%, respectively. Inhibition of the cytochrome P450 monooxygenase completely abolished the effect of the extracellular Ca(2)+ on channel activity in the mTAL from rats on a different K diet. In contrast, blocking cyclooxygenase did not significantly alter the responsiveness of the 70-pS K channel to the extracellular Ca(2)+. Moreover, addition of sodium nitropruside, a nitric oxide (NO) donor, not only increased the channel activity, but also blunted the inhibitory effect of the extracellular Ca(2)+ on the 70-pS K channel and decreased 20-hydroxyeicosatetraenoic acid (20-HETE) concentration in the mTAL from rats on a KD diet. In contrast, inhibiting NOS with L-NAME enhanced the inhibitory effect of the extracellular Ca(2)+ on the channel activity and increased 20-HETE concentration in the mTAL from rats on a high K diet. Western blot has further shown that the expression of inducible NO synthase (iNOS) is significantly higher in the renal medulla from rats on an HK diet than that on a KD diet. Also, addition of S-nitroso-N-acetylpenicillamine abolished the inhibitory effect of arachidonic acid on channel activity in the mTAL, whereas it did not block the inhibitory effect of 20-HETE. We conclude that a low dietary K intake increases the sensitivity of the 70-pS K channel to the extracellular Ca(2)+, and that a decrease in NOS activity is involved in enhancing the inhibitory effect of the extracellular Ca(2)+ on channel activity in the mTAL during K depletion.

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A channel recording showing the effect of 5 μM arachidonic acid on the activity of the 70-pS K channel in the presence of 10 μM SNAP. The experiment was performed in a cell-attached patch in the mTAL from a rat on a high K diet, and the pipette holding potential was 0 mV. The top trace shows the time course, and three parts of the trace are extended to demonstrate the fast time resolution.
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Figure 11: A channel recording showing the effect of 5 μM arachidonic acid on the activity of the 70-pS K channel in the presence of 10 μM SNAP. The experiment was performed in a cell-attached patch in the mTAL from a rat on a high K diet, and the pipette holding potential was 0 mV. The top trace shows the time course, and three parts of the trace are extended to demonstrate the fast time resolution.

Mentions: The notion that a decrease in NOS activity may be partially responsible for increasing the 20-HETE generation is also supported by the finding that the 20-HETE production was diminished significantly from 8.2 ± 1 to 5.8 ± 1 pg/μg protein (n = 5 rats) in the mTALs treated with 20 μM SNP from rats on a KD diet (Fig. 10). In contrast, treatment of the mTAL with 0.2 mM L-NAME increased the 20-HETE production from 2.2 ± 0.3 to 4.1 ± 0.5 pg/μg protein (n = 4 rats) in the tubules harvested from rats on a high K diet. To further demonstrate that increasing NO concentration could attenuate the inhibitory effect of AA, we examined the effect of AA in the presence of NO. We confirmed the previous finding (Wang and Lu 1995) that application of 5 μM AA inhibited the activity of the 70-pS K channel and decreased NPo by 80 ± 6% (n = 5 patches). Moreover, application of 10 μM SNAP, a donor of NO, completely reversed the inhibitory effect of AA and increased the NPo from 0.10 ± 0.02 to 1.1 ± 0.1 (n = 4; Fig. 11). The effect of SNAP is not the result of a direct stimulation of channel activity since addition of SNAP had no effect on channel activity in inside-out patches (unpublished data). Furthermore, the notion that the effect of SNAP on the AA-induced inhibition of channel activity results from suppressing the cytochrome P450 metabolism of AA was also confirmed by the finding that SNAP did not abolish the inhibitory effect of 20-HETE on channel activity in inside-out patches or cell-attached patches. Fig. 12 is a recording made in a cell-attached patch showing that application of 100 nM 20-HETE inhibited the 70-pS K channel by 90 ± 8% (n = 4 patches) in the presence of SNAP. Moreover, we confirmed the previous finding that the sensitivity of the 70-pS K channel to 20-HETE is the same in the mTAL from rats on a different K diet (Gu et al. 2001). Therefore, the results suggest that the effect of NO takes place before 20-HETE formation.


K depletion enhances the extracellular Ca2+-induced inhibition of the apical K channels in the mTAL of rat kidney.

Gu RM, Wei Y, Jiang HL, Lin DH, Sterling H, Bloom P, Balazy M, Wang WH - J. Gen. Physiol. (2002)

A channel recording showing the effect of 5 μM arachidonic acid on the activity of the 70-pS K channel in the presence of 10 μM SNAP. The experiment was performed in a cell-attached patch in the mTAL from a rat on a high K diet, and the pipette holding potential was 0 mV. The top trace shows the time course, and three parts of the trace are extended to demonstrate the fast time resolution.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2233853&req=5

Figure 11: A channel recording showing the effect of 5 μM arachidonic acid on the activity of the 70-pS K channel in the presence of 10 μM SNAP. The experiment was performed in a cell-attached patch in the mTAL from a rat on a high K diet, and the pipette holding potential was 0 mV. The top trace shows the time course, and three parts of the trace are extended to demonstrate the fast time resolution.
Mentions: The notion that a decrease in NOS activity may be partially responsible for increasing the 20-HETE generation is also supported by the finding that the 20-HETE production was diminished significantly from 8.2 ± 1 to 5.8 ± 1 pg/μg protein (n = 5 rats) in the mTALs treated with 20 μM SNP from rats on a KD diet (Fig. 10). In contrast, treatment of the mTAL with 0.2 mM L-NAME increased the 20-HETE production from 2.2 ± 0.3 to 4.1 ± 0.5 pg/μg protein (n = 4 rats) in the tubules harvested from rats on a high K diet. To further demonstrate that increasing NO concentration could attenuate the inhibitory effect of AA, we examined the effect of AA in the presence of NO. We confirmed the previous finding (Wang and Lu 1995) that application of 5 μM AA inhibited the activity of the 70-pS K channel and decreased NPo by 80 ± 6% (n = 5 patches). Moreover, application of 10 μM SNAP, a donor of NO, completely reversed the inhibitory effect of AA and increased the NPo from 0.10 ± 0.02 to 1.1 ± 0.1 (n = 4; Fig. 11). The effect of SNAP is not the result of a direct stimulation of channel activity since addition of SNAP had no effect on channel activity in inside-out patches (unpublished data). Furthermore, the notion that the effect of SNAP on the AA-induced inhibition of channel activity results from suppressing the cytochrome P450 metabolism of AA was also confirmed by the finding that SNAP did not abolish the inhibitory effect of 20-HETE on channel activity in inside-out patches or cell-attached patches. Fig. 12 is a recording made in a cell-attached patch showing that application of 100 nM 20-HETE inhibited the 70-pS K channel by 90 ± 8% (n = 4 patches) in the presence of SNAP. Moreover, we confirmed the previous finding that the sensitivity of the 70-pS K channel to 20-HETE is the same in the mTAL from rats on a different K diet (Gu et al. 2001). Therefore, the results suggest that the effect of NO takes place before 20-HETE formation.

Bottom Line: Physiol. 270:C103-C111).Also, addition of S-nitroso-N-acetylpenicillamine abolished the inhibitory effect of arachidonic acid on channel activity in the mTAL, whereas it did not block the inhibitory effect of 20-HETE.We conclude that a low dietary K intake increases the sensitivity of the 70-pS K channel to the extracellular Ca(2)+, and that a decrease in NOS activity is involved in enhancing the inhibitory effect of the extracellular Ca(2)+ on channel activity in the mTAL during K depletion.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, New York Medical College, Valhalla, NY 10595, USA.

ABSTRACT
We have shown previously that raising extracellular Ca(2)+ inhibited the apical 70-pS K channel in the thick ascending limb (TAL; Wang, W.H., M. Lu, and S.C. Hebert. 1996. Am. J. Physiol. 270:C103-C111). We now used the patch-clamp technique to study the effect of increasing the extracellular Ca(2)+ on the 70-pS K channel in the mTAL from rats on a different K diet. Increasing the extracellular Ca(2)+ from 10 microM to 0.5, 1, and to 1.5 mM in the mTAL from rats on a K-deficient (KD) diet inhibited the channel activity by 30, 65, and 90%, respectively. In contrast, raising the extracellular Ca(2)+ to 1.5 mM had no significant effect on channel activity in the mTAL from animals on a high K (HK) diet and further increasing the extracellular Ca(2)+ to 2.5, 3.5, and 5.5 mM decreased the channel activity by 29, 55, and 90%, respectively. Inhibition of the cytochrome P450 monooxygenase completely abolished the effect of the extracellular Ca(2)+ on channel activity in the mTAL from rats on a different K diet. In contrast, blocking cyclooxygenase did not significantly alter the responsiveness of the 70-pS K channel to the extracellular Ca(2)+. Moreover, addition of sodium nitropruside, a nitric oxide (NO) donor, not only increased the channel activity, but also blunted the inhibitory effect of the extracellular Ca(2)+ on the 70-pS K channel and decreased 20-hydroxyeicosatetraenoic acid (20-HETE) concentration in the mTAL from rats on a KD diet. In contrast, inhibiting NOS with L-NAME enhanced the inhibitory effect of the extracellular Ca(2)+ on the channel activity and increased 20-HETE concentration in the mTAL from rats on a high K diet. Western blot has further shown that the expression of inducible NO synthase (iNOS) is significantly higher in the renal medulla from rats on an HK diet than that on a KD diet. Also, addition of S-nitroso-N-acetylpenicillamine abolished the inhibitory effect of arachidonic acid on channel activity in the mTAL, whereas it did not block the inhibitory effect of 20-HETE. We conclude that a low dietary K intake increases the sensitivity of the 70-pS K channel to the extracellular Ca(2)+, and that a decrease in NOS activity is involved in enhancing the inhibitory effect of the extracellular Ca(2)+ on channel activity in the mTAL during K depletion.

Show MeSH
Related in: MedlinePlus