Vasopressin and PGE2 regulate activity of apical 70 pS K+ channel in thick ascending limb of rat kidney

Hua Jun Liu, Yuan Wei, Nicholas R. Fererri, Alberto Nasjletti, and Wen Hui Wang

Department of Pharmacology, New York Medical College, Valhalla, New York 10595


    ABSTRACT
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Vasopressin and prostaglandin E2 (PGE2) are involved in regulating NaCl reabsorption in the thick ascending limb (TAL) of the rat kidney. In the present study, we used the patch-clamp technique to study the effects of vasopressin and PGE2 on the apical 70 pS K+ channel in the rat TAL. Addition of vasopressin increased the channel activity, defined as NPo, from 1.11 to 1.52 (200 pM) and 1.80 (500 pM), respectively. The effect of vasopressin can be mimicked by either forskolin (1-5 µM) or 8-bromo-cAMP/dibutyryl-cAMP (8-Br-cAMP/DBcAMP) (200-500 µM). Moreover, the effects of cAMP and vasopressin were not additive and application of 10 µM H-89 abolished the effect of vasopressin. This suggests that the effect of vasopressin is mediated by a cAMP-dependent pathway. Applying 10 nM PGE2 alone had no significant effect on the channel activity. However, PGE2 (10 nM) abolished the stimulatory effect of vasopressin. The PGE2-induced inhibition of the vasopressin effect was the result of decreasing cAMP production because addition of 200 µM 8-Br-cAMP/DBcAMP reversed the PGE2-induced inhibition. In addition to antagonizing the vasopressin effect, high concentrations of PGE2 reduced channel activity in the absence of vasopressin by 33% (500 nM) and 51% (1 µM), respectively. The inhibitory effect of high concentrations of PGE2 was not the result of decreasing cAMP production because adding the membrane-permeant cAMP analog failed to restore the channel activity. In contrast, inhibiting protein kinase C (PKC) with calphostin C (100 nM) abolished the effect of 1 µM PGE2. We conclude that PGE2 inhibits apical K+ channels by two mechanisms: 1) low concentrations of PGE2 attenuate the vasopressin-induced stimulation mainly by reducing cAMP generation, and 2) high concentrations of PGE2 inhibit the channel activity by a PKC-dependent pathway.

cyclooxygenase; cAMP; protein kinase C; patch-clamp technique; NaCl transport


    INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

THE THICK ASCENDING LIMB (TAL) is responsible for reabsorption of 20-25% of the filtrated Na+ load and plays a key role in the urinary concentrating mechanism (16, 17). Na+ enters the cell through the luminal Na+-K+-Cl- cotransporter energized by a favorable electrochemical gradient of Na+ and is then pumped out of the cell through the basolateral Na+-K+-ATPase. K+ recycling is essential for maintaining the normal function of the Na+-K+-Cl- cotransporter (16, 19). First, K+ recycling hyperpolarizes the cell membrane and accordingly provides the driving force for Cl- leaving the cell. Second, K+ recycling participates in generating the lumen-positive potential, which is the driving force for transepithelial Na+, Ca2+, and Mg2+ transport. Third, K+ recycling is important for providing an adequate supply of K+ to the Na+-K+-Cl- cotransporter in the cortical TAL where the K+ concentration is at least one order of magnitude lower than that of Na+ and Cl-.

Three types of K+ channels, 70 pS, 30 pS, and Ca2+-activated maxi K+ (>100 pS), have been expressed in the apical membrane of the TAL (5, 32, 33). However, it is generally believed that the 70 pS and the 30 pS K+ channels are responsible for K+ recycling across the apical membrane (34). This conclusion is based on observations that both channels have a higher channel open probability as well as a larger number of channels per cell than that of the maxi K+ channel in the TAL (5, 33). The 70 pS K+ channel is sensitive to MgATP, inhibited by acidic pH and protein kinase C (PKC), and stimulated by cAMP-dependent protein kinase (PKA) (5, 35). Furthermore, patch-clamp experiments demonstrated that the 70 pS K+ channel and the 30 pS K+ channel contribute to the apical K+ conductance by ~80% and 20%, respectively (35). Thus the 70 pS K+ channel should play a key role in K+ recycling.

Prostaglandin E2 (PGE2) plays an important role in regulating Cl- reabsorption in the TAL (13, 21, 27). The effect of PGE2 is mediated by inhibition of vasopressin-induced increase in cAMP level. Because K+ recycling plays an important role in maintaining the function of the Na+-K+-Cl- cotransporter, it is possible that the effect of PGE2 on NaCl transport in the TAL involves the regulation of apical K+ channels. Therefore, the present study is designed to investigate the effect of interaction between PGE2 and vasopressin on channel activity.


    METHODS
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Preparation of the TAL. The pathogen-free Sprague-Dawley rats (100-120 g) used in experiments were purchased from Taconic (Germantown, NY). Animals were killed by cervical dislocation after anesthesia with Metofane. The kidneys were removed immediately and thin coronal sections were cut with a razor blade. We used only medullary TALs in the study. The dissection buffer solution contained (in mM) 140 NaCl, 5 KCl, 1.5 MgCl2, 1.8 CaCl2, 5 glucose, and 10 HEPES (pH 7.4 with NaOH) at 22°C. The isolated tubule was transferred onto a 5 × 5 mm cover glass coated with Cell-Tak (Collaborative Research, Bedford, MA) to immobilize the tubule. The cover glass was placed in a chamber mounted on an inverted microscope (Nikon) and the tubules were superfused with HEPES-buffered NaCl solution, composed of (in mM) 140 NaCl, 5 KCl, 1.8 CaCl2, 1.8 MgCl2, and 10 HEPES (pH 7.4). The TAL was cut open with a sharpened micropipette to gain access to the apical membrane. The temperature of the chamber (1,000 µl) was maintained at 37 ± 1°C by circulating warm water around the chamber.

Patch-clamp technique. We used an Axon200A patch-clamp amplifier to record channel current. The current was low-pass filtered at 1 kHz using an eight-pole Bessel filter (902LPF; Frequency Devices, Haverhill, MA) and was digitized at a sampling rate of 44 kHz using a digital data recorder (model VR-10B; Instrutech Great Neck, NY) and stored on videotape (Sony SL-2700). For analysis, data stored on the tape was collected to an IBM-compatible Pentium computer (Gateway 2000) at a rate of 4 kHz and analyzed using the pCLAMP software system 6.04 (Axon Instruments, Burlingame, CA). Opening and closing transitions were detected using 50% of the single-channel amplitude as the threshold. Channel activity was defined as NPo. The NPo was calculated from data samples of 30-60 s duration that were always at the end of each experimental maneuver and in the steady state. We used the following equation to obtain NPo
<IT>NP</IT><SUB>o</SUB> = &Sgr;(<IT>t</IT><SUB>1</SUB> + <IT>t</IT><SUB>2</SUB> + … <IT>t<SUB>i</SUB></IT>) (1)
where ti is the fractional open time spent at each of the observed current levels. Because two types of K+ channels are present in the apical membrane of TAL, two types of K+ channels sometimes can be detected in the same patches. To avoid this complexity, we have selected those patches with only a single population of K+ channel to calculate NPo. If the low-conductance K+ channel appeared during the course of experiments, we used a ruler to calculate NPo manually.

Chemicals and experimental solution. The pipette solution contained (in mM) 140 KCl, 1.8 MgCl2, and 10 HEPES (pH=7.4). Forskolin, 8-Br-cAMP, and DBcAMP were purchased from Sigma (St. Louis, MO) and forskolin was dissolved in DMSO. PGE2 and calphostin C were obtained from Biomol (Plymouth Meeting, PA) and dissolved in ethanol. The final concentration of either DMSO or ethanol in experiments was less than 0.1% and had no effect on channel activity. The chemicals were added directly to the bath to reach the final concentration.

Statistics. Data are shown as mean ± SE. We used paired Student's t-tests to determine the significance of difference between the control and experimental periods. Statistical significance was taken as P < 0.05.


    RESULTS
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

We have previously observed that application of vasopressin stimulated the 30 pS K+ channel in the apical membrane of the TAL (33). We extended our study to examine the effect of vasopressin on the activity of the 70 pS K+ channel. Figure 1A is a representative channel recording showing the effect of vasopressin on the 70 pS K+ channel in a cell-attached patch. It is apparent that vasopressin significantly stimulates the activity of the 70 pS K+ channel. Application of 200 and 500 pM vasopressin increased the NPo from 1.11 ± 0.1 to 1.52 ± 0.16 and 1.80 ± 0.2 (n = 7), respectively (Fig. 1B). The effect of vasopressin was reversible because the channel activity returned to the control value after removal of vasopressin. To determine whether the effect of vasopressin is mediated by a cAMP-dependent pathway, we investigated the effect of 1-5 µM forskolin, an agent that stimulates the adenylate cyclase, on channel activity. Figure 2A is a representative recording demonstrating the effect of forskolin on the 70 pS K+ channel in a cell-attached patch. In five experiments, addition of 1-5 µM forskolin increased the NPo from 1.16 ± 0.1 to 1.85 ± 0.2, and the effect was reversible as the channel activity returned to the control value after removal of forskolin. That the effect of vasopressin was mediated by a cAMP-dependent pathway is further suggested by observations that application of the membrane-permeant cAMP analogs stimulates the channel activity. Figure 2B is a typical recording illustrating the effect of 8-Br-cAMP/DBcAMP on channel activity in a cell-attached patch. Addition of 200-500 µM 8-Br-cAMP/DBcAMP increased NPo from 0.95 ± 0.1 to 1.4 ± 0.12 (n = 4). Figure 2C summarizes the results regarding the effect of vasopressin, 8-Br-cAMP/DBcAMP, and forskolin. Vasopressin (500 pM) significantly increased channel activity by 62 ± 7% (n = 7). Similarly, addition of 500 µM 8-Br-cAMP/DBcAMP or 1-5 µM forskolin significantly raised channel activity by 47 ± 5% (n = 4) and 59 ± 5% (n = 5), respectively. Moreover, the effects of vasopressin and cAMP were not additive because addition of vasopressin (500 pM) + DBcAMP increased channel activity by 63 ± 7% (n = 4), a value that is not significantly different from that observed with vasopressin alone. This indicates that the effect of vasopressin is the result of stimulating cAMP generation. To determine whether the effect of vasopressin is the result of inhibition of PKA, we examined the effect of vasopressin in the presence of H-89, an inhibitor of PKA (Fig. 3). Addition of H-89 (10 µM) had no significant effect on channel activity (control NPo = 0.95 ± 0.08, H-89 = 0.85 ± 0.08). This suggests that PKA is not required in maintaining the activity of the opened channels. However, inhibition of PKA completely abolished the effect of vasopressin because it failed to increase NPo (0.82 ± 0.09) in the presence of a PKA inhibitor (n = 3). This indicates that the effect of vasopressin is mediated by PKA and is most likely the result of stimulation of a V2 cAMP-coupled receptor.


View larger version (22K):
[in this window]
[in a new window]
 
Fig. 1.   A: a channel recording shows effect of vasopressin (AVP) on activity of 70 pS K+ channel in apical membrane of mTAL. Experiment was performed in a cell-attached patch and holding potential was 0 mV. C, channel closed level. B: effect of AVP on activity of apical 70 pS K+ channel. * Data significantly different from control value; NPo, channel activity.



View larger version (26K):
[in this window]
[in a new window]
 
Fig. 2.   A: channel recording showing effect of 1 µM forskolin. Experiment was carried out in a cell-attached patch and holding potential was 0 mV. B: recording demonstrates effect of 0.5 mM DBcAMP on channel activity. Experiment was performed in a cell-attached patch and holding potential was -30 mV. C: effects of 500 pM AVP, 500 µM DBcAMP/8-Br-cAMP (cAMP), 1-5 µM forskolin, and cAMP + AVP on channel activity in cell-attached patches. Data are significantly different from control value.



View larger version (20K):
[in this window]
[in a new window]
 
Fig. 3.   A: channel recording showing effect of AVP in presence of 10 µM H-89. Experiment was carried out in a cell-attached patch and holding potential was 0 mV. B: effect of 10 µM H-89 and AVP (500 pM) + 10 µM H-89 on channel activity.

In addition to a V2 receptor, PGE2 type III receptor, EP3, is present in the TAL and stimulating the EP3 receptor has been shown to reduce cAMP production (2, 25). Thus we explored the interaction between vasopressin and the effect of PGE2 to determine whether PGE2 can abolish the effect of vasopressin. Figure 4A is a recording showing the effect of PGE2 on the vasopressin-induced increase in channel activity. It is clear that 10 nM PGE2 abolished the vasopressin-induced increase in channel activity. The inhibitory effect of PGE2 resulted from blocking vasopressin-induced increase in cAMP concentration because addition of cAMP analogs reversed the effect of PGE2. Figure 4B summarizes the results from five experiments that examined the interaction between vasopressin and PGE2. Adding 200 pM vasopressin increased NPo from the control value (0.80 ± 0.07) to 1.14 ± 0.10. However, in the presence of vasopressin, application of 10 nM PGE2 abolished the effect of vasopressin and the NPo decreased to 0.75 ± 0.07, a value that is not significantly different from the control. Moreover, addition of 200 µM DBcAMP raised the NPo from 0.75 ± 0.07 to 1.07 ± 0.10 in the presence of PGE2.


View larger version (19K):
[in this window]
[in a new window]
 
Fig. 4.   A: channel recording shows that prostaglandin E2 (PGE2) abolished stimulatory effect of 200 pM AVP. Experiment was performed in a cell-attached patch and holding potential was 0 mV. Arrows indicate where PGE2 and DBcAMP (200 µM) were added. B: effects of AVP (200 µM), PGE2 (10 nM) + AVP, and 200 µM DBcAMP/8-Br-cAMP + PGE2 + AVP on channel activity. *Data significantly different from control.

That the effect of 10 nM PGE2 was the result of inhibiting vasopressin-induced increase in cAMP level was further confirmed by experiments where in the absence vasopressin, 10 nM PGE2 had no significant effect on the channel activity (Fig. 5A). However, further increase of PGE2 concentration to 1 µM significantly decreased channel activity by 51%, and NPo decreased from 1.39 to 0.67. The inhibition induced by high concentrations of PGE2 was not the result of decreasing cAMP concentration because adding 500 µM DBcAMP failed to restore the channel activity and NPo slightly increased from 0.67 to 0.8 (Fig. 5B).


View larger version (23K):
[in this window]
[in a new window]
 
Fig. 5.   A: channel recording shows effect of 10 nM PGE2, 1 µM PGE2, and 500 µM DBcAMP + 1 µM PGE2. Experiment was carried out in a cell-attached patch and holding potential was -30 mV. B: dose-response curve of PGE2 on channel activity. Experiments were performed in absence of AVP. *Significantly different from control (100%).

Figure 5B is a dose-response curve of PGE2 effect in the absence of vasopressin, showing that addition of 500 nM, 1 µM, and 10 µM PGE2 reduced channel activity by 33 ± 3% (n = 4), 51 ± 5% (n = 11), and 55 ± 5% (n = 4), respectively. Because PGE2 has been shown to stimulate PKC, we investigated the role of PKC in mediating the effect of high concentrations of PGE2. Figure 6 is a representative recording from seven such experiments showing the effect of calphostin C, an agent that inhibits PKC, on 1 µM PGE2-induced decrease in channel activity. As demonstrated in Fig. 6A, 1 µM PGE2 reduced channel activity by 51 ± 5%, and adding 500 µM DBcAMP increased channel activity slightly to 60 ± 7% of the control value. In contrast, inhibiting PKC completely restored channel activity to the control value (Fig. 6B). The notion that the effect of high concentrations of PGE2 is mediated by a PKC-dependent pathway was also indicated by experiments where in the presence of calphostin C, the inhibitory effect of 1 µM PGE2 was completely abolished (Figs. 6 and 7). Figure 7 is a representative recording from five experiments showing that addition of calphostin C had no significant effect on channel activity (105 ± 5% of the control). However, in the presence of a PKC inhibitor, addition of 1 µM PGE2 had no significant effect on the K+ channel because channel activity was still 102 ± 5% of the control value.


View larger version (23K):
[in this window]
[in a new window]
 
Fig. 6.   A: channel recording demonstrates effect of 1 µM PGE2 on channel activity in presence of 500 µM DBcAMP and 100 nM calphostin C. Experiment was performed in a cell-attached patch and holding potential was 0 mV. B: effects of 10 nM PGE2, 1 µM PGE2, 500 µM DBcAMP + 1 µM PGE2, and 100 nM calphostin C + 1 µM PGE2 on channel activity. Channel activity was normalized by comparing to corresponding control NPo. * Data significantly different from control value.



View larger version (50K):
[in this window]
[in a new window]
 
Fig. 7.   Channel recording showing effect of 1 µM PGE2 in presence of 100 nM calphostin C. Experiment was performed in a cell-attached patch and holding potential was 0 mV. Time course of experiments (A) and 3 parts of trace (B) are demonstrated at a fast time resolution.


    DISCUSSION
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

The main finding of the present study is that low concentrations of PGE2 abolish the stimulatory effect of vasopressin on the apical 70 pS K+ channel, whereas high concentrations of PGE2 inhibit the channel activity via a PKC-dependent pathway. Because the 70 pS K+ channel is the most abundant K+ channel in the apical membrane of the rat TAL, the effect of PGE2 on the 70 pS K+ channel should inhibit the K+ recycling across the apical membrane.

That K+ recycling is essential for maintaining normal function of the Na+-K+-Cl- cotransporter is convincingly demonstrated by a genetic study showing that a defective gene product encoding renal outer medulla K+ (ROMK) channels led to an abnormal salt transport in human kidney, Bartter's syndrome (30). Although it is well established that the ROMK channel family is a key component of the 30 pS K+ channel (26, 34), the relationship between the 70 pS K+ channel and ROMK is not clear. Because the 70 pS K+ channel could contribute as much as 80% of K+ conductance to the apical membrane of the rat TAL, K+ recycling is unlikely to be severely compromised by only a 20% decrease in the apical K+ conductance. Thus it is possible that ROMK is also involved in forming the 70 pS K+ channel. If the 70 pS K+ channel is not expressed in the apical membrane of the TAL of human kidney, the mechanism by which PGE2 inhibits the 70 pS K+ channel is still physiologically relevant for understanding the role of PGE2 in regulating K+ recycling across the apical membrane of the TAL because ROMK channels are also regulated by PKA and PKC (24, 34, 37).

In the present study, we have demonstrated that vasopressin activates the apical K+ channels. Vasopressin has been shown to stimulate NaCl transport in the TAL of mouse kidney (17). The effect of vasopressin is mediated by a cAMP-dependent pathway because cAMP mimics the effect of vasopressin. Moreover, the effect is the result of stimulating PKA-induced phosphorylation of basolateral Cl- channels, apical Na+-K+-Cl- cotransporters, and K+ channels (16, 18, 20). In previous studies, we demonstrated that vasopressin stimulated the 30 pS K+ channel in the rat TAL and cortical collecting duct (11, 33). Now, we have shown that vasopressin can also increase the activity of the 70 pS K+ channel. Three lines of evidence support the notion that the effect of vasopressin is mediated by a cAMP-dependent pathway: 1) addition of either forskolin or membrane-permeant cAMP analogs mimics the effect of vasopressin, 2) the effects of vasopressin and cAMP are not additive, and 3) inhibition of PKA abolishes the effect of vasopressin. The notion that PKA mediates the stimulatory effect of vasopressin is also confirmed by studies on renal medullary vesicles (28).

In addition to vasopressin, PGE2 has been shown to play an important role in regulating NaCl transport in the TAL (12, 13, 21, 31). Several studies have demonstrated that PGE2 blocks the stimulatory effect of vasopressin on NaCl transport in the TAL (12, 13, 21). Moreover, the effect of PGE2 was induced by inhibiting vasopressin-dependent increase in cAMP generation (7, 13). Although we cannot completely exclude the role of PKC, two lines of evidence indicate that the effect of low concentrations (<100 nM) of PGE2 on channel activity is mainly produced by decreasing vasopressin-induced increase in cAMP generation. First, PGE2 had no significant effect on channel activity in the absence of vasopressin. Second, applying exogenous cAMP analogs abolishes the inhibitory effect of PGE2. In addition to antagonizing the effect of vasopressin, we demonstrated that high concentrations of PGE2 (>500 nM) can inhibit channel activity via a cAMP-independent mechanism. Because blocking PKC abolishes the inhibition induced by high concentrations of PGE2, the effect is most likely mediated by a PKC-dependent pathway.

Four types of PGE2 receptors, EP1, EP2, EP3, and EP4, have been identified in kidneys (1, 4, 7, 8, 36). Immunocytochemical studies have revealed that the main type of PGE2 receptor expressed in the mTAL is the EP3 receptor, which is coupled to Gi protein. Thus the effect of low concentrations of PGE2 should be the result of stimulating the Gi-coupled EP3 receptor. However, the EP1 receptor, which is linked to phospholipase C, is not identified in the mTAL (8, 9). There are two possibilities to explain how high concentrations of PGE2 can stimulate the PKC-dependent pathway: 1) some EP3 receptors with low affinity could be alternatively coupled to phopholipase C, and 2) a low level of EP1 receptors, which is not detected with the current methodology, may be expressed in the mTAL. The first possibility is supported by findings showing that EP3 receptors have several alternatively spliced variants and these variants can couple to different signaling mechanisms including phosphatidylinositol biphosphate hydrolysis (2, 25). Because the EP3 receptor, which does not couple to Gi protein, has a low affinity to PGE2, low concentrations of PGE2 cannot activate these variants. Our finding that PGE2 could activate the PKC-dependent pathway is also consistent with Good's observation that PGE2 causes selective activation of the PKC-delta isoform (3, 15). Moreover, the fact that high concentrations of PGE2 inhibit the apical K+ channels by a PKC-dependent mechanism may also be important to understand the beneficial effect of inhibiting prostaglandin synthesis in treatment of Bartter's syndrome. One of the characteristics of Bartter's syndrome is an increase in plasma PGE2 level (29). Inhibition of cyclooxygenase (COX) improves the manifestation of the disease (27). Because NaCl transport in the TAL is severely compromised in patients with Bartter's syndrome, high concentrations of PGE2 further diminish the remaining function of the TAL. Therefore, inhibition of PGE2 production should improve the transport function of the TAL cells.

PGE2 is the product of COX-dependent metabolism of arachidonic acid (AA). There are at least two AA metabolic pathways in the mTAL: COX and cytochrome P-450 monooxygenase-dependent metabolisms (6, 23). A major metabolite of the cytochrome P-450-dependent pathway in the mTAL (10), 20-hydroxyeicosatetraenoic acid, has been shown to regulate the activity of the 70 pS K+ channel (35) and Na+-K+-Cl- cotransporter (14). Thus the cytochrome P-450-dependent pathway should play a key role in the regulation of NaCl transport in the TAL (34). On the other hand, hypercalcemia has been shown to increase PGE2 production. This increase in PGE2 may be partially responsible for diminished transport function in the loop of Henle during hypercalcemia (22). However, it is not known whether the COX-dependent pathway interacts with P-450-dependent metabolism of AA. We need further experiments to explore the role of interaction between the two AA metabolic pathways in the regulation of the transport function of the TAL.

Figure 8 is a model of the TAL cell illustrating the mechanism by which PGE2 regulates the apical K+ channels and possibly the NaCl transport in the mTAL. Low concentrations of PGE2 antagonize the stimulatory effect of vasopressin on the apical K+ conductance, whereas high concentrations inhibit the apical K+ channel by PKC stimulation.


View larger version (22K):
[in this window]
[in a new window]
 
Fig. 8.   Model of thick ascending limb (TAL) cell illustrating mechanism by which PGE2 regulates apical K+ channels. Arrows and double bars represent stimulation and inhibition, respectively; PKA, cAMP-dependent protein kinase; PKC, protein kinase C; AC, adenylyl cyclase.


    ACKNOWLEDGEMENTS

We thank M. Steinberg for editorial assistance.


    FOOTNOTES

This work was supported by National Heart, Lung, and Blood Institute Grant PO1HL-34300 and National Institute of Diabetes and Digestive and Kidney Diseases Grant DK-47402.

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

Address for reprint requests and other correspondence: W. H. Wang, Dept. of Pharmacology, New York Medical College, Grasslands Reservation, Valhalla, NY 10595 (E-mail: wenhui_wang{at}nymc.edu).

Received 12 August 1999; accepted in final form 17 November 1999.


    REFERENCES
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

1.   Adam, M, Boie Y, Rushmore TH, Müller G, Bastien L, McKee KT, Metters KM, and Abramovitz M. Cloning and expression of three isoforms of the human EP3 prostanoid receptor. FEBS Lett 338: 170-174, 1994[ISI][Medline].

2.   An, Y, Yang J, So S, Zeng L, and Goetzl E. Isoforms of the EP3 subtype of human PGE2 receptor transduce both intracellular calcium and cAMP signals. Biochemistry 33: 14496-14502, 1994[ISI][Medline].

3.   Aristimuno, P, and Good DW. PKC isoforms in rat medullary thick ascending limb: selective activation of the delta -isoform by PGE2. Am J Physiol Renal Physiol 272: F624-F631, 1997[Abstract/Free Full Text].

4.   Bastien, L, Sawyer N, Grygorczyk R, Metters KM, and Adam M. Cloning, functional expression, and characterization of the human prostaglandin E2 receptor EP2 subtype. J Biol Chem 269: 11873-11877, 1994[Abstract/Free Full Text].

5.   Bleich, M, Schlatter E, and Greger R. The luminal K+ channel of the thick ascending limb of Henle's loop. Pflügers Arch 415: 449-460, 1990[ISI][Medline].

6.   Bonvalet, JP, Pradelles P, and Farman N. Segmental synthesis and actions of prostaglandins along the nephron. Am J Physiol Renal Fluid Electrolyte Physiol 253: F377-F387, 1987[Abstract/Free Full Text].

7.   Breyer, MD, and Ando Y. Hormonal signaling and regulation of salt and water transport in the collecting duct. Annu Rev Physiol 56: 711-739, 1994[ISI][Medline].

8.   Breyer, MD, Davis L, Jacobson HR, and Breyer RM. Differential localization of prostaglandin E receptor subtypes in human kidney. Am J Physiol Renal Fluid Electrolyte Physiol 270: F912-F918, 1996[Abstract/Free Full Text].

9.   Breyer, MD, Jacobson HR, Davis LS, and Breyer RM. In situ hybridization and localization of mRNA for the rabbit prostaglandin EP3 receptor. Kidney Int 43: 1372-1378, 1993.

10.   Carroll, MA, Sala A, Dunn CE, McGiff JC, and Murphy RC. Structural identification of cytochrome P-450-dependent arachidonate metabolites formed by rabbit medullary thick ascending limb cells. J Biol Chem 266: 12306-12312, 1991[Abstract/Free Full Text].

11.   Cassola, AC, Giebisch G, and Wang WH. Vasopressin increases density of apical low-conductance K+ channels in rat CCD. Am J Physiol Renal Fluid Electrolyte Physiol 264: F502-F509, 1993[Abstract/Free Full Text].

12.   Culpepper, RM, and Andreoli TE. Interactions among prostaglandin E2, antidiuretic hormone, and cyclic adenosine monophosphate in modulating Cl- absorption in single mouse medullary thick ascending limbs of Henle. J Clin Invest 71: 1588-1601, 1983[ISI][Medline].

13.   Culpepper, RM, and Andreoli TE. Prostaglandin E2 inhibition of vasopressin-stimulated NaCl transport in the mouse medullary thick ascending limb of Henle. In: Prostaglandins and Membrane Ion Transport, edited by Braquet P.. New York: Raven, 1984, p. 327-334.

14.   Escalante, B, Erlij D, Falck JR, and McGiff JC. Effect of cytochrome P-450 arachidonate metabolites on ion transport in rabbit kidney loop of Henle. Science 251: 799-802, 1991[ISI][Medline].

15.   Good, DW. PGE2 reverses AVP inhibition of HCO-3 absorption in rat mTAL by activation of protein kinase C. Am J Physiol Renal Fluid Electrolyte Physiol 270: F978-F985, 1996[Abstract/Free Full Text].

16.   Greger, R. Ion transport mechanisms in thick ascending limb of Henle's loop of mammalian nephron. Physiol Rev 65: 760-797, 1985[Free Full Text].

17.   Hebert, SC, and Andreoli TE. Control of NaCl transport in the thick ascending limb. Am J Physiol Renal Fluid Electrolyte Physiol 246: F745-F756, 1984[Abstract/Free Full Text].

18.   Hebert, SC, and Andreoli TE. Effects of antidiuretic hormone on cellular conductive pathways in mouse medullary thick ascending limbs of Henle. II. Determinants of the ADH-mediated increases in transepithelial voltage and in net Cl absorption. J Membr Biol 80: 221-233, 1984[ISI][Medline].

19.   Hebert, SC, Culpepper RM, and Andreoli TE. NaCl transport in mouse medullary thick ascending limbs. II. ADH enhancement of transcellular NaCl cotransport; origin of transepithelial voltage. Am J Physiol Renal Fluid Electrolyte Physiol 241: F432-F442, 1981[Abstract/Free Full Text].

20.   Hebert, SC, Friedman PA, and Andreoli TE. Effects of antidiuretic hormone on cellular conductive pathways in mouse medullary thick ascending limb of Henle. I. ADH increases transcellular conductance pathways. J Membr Biol 80: 201-219, 1984[ISI][Medline].

21.   Kaji, DM, Chase HS, Jr, Eng JP, and Diaz J. Prostaglandin E2 inhibits Na-K-2Cl cotransport in medullary thick ascending limb cells. Am J Physiol Renal Fluid Electrolyte Physiol 271: F354-F361, 1996.

22.   Levi, M, Peterson L, and Berl T. Mechanism of concentrating defect in hypercalcemia. Role of polydipsia and prostaglandins. Kidney Int 23: 489-497, 1983[ISI][Medline].

23.   McGiff, JC. Cytochrome P-450 metabolism of arachidonic acid. Annu Rev Pharmacol Toxicol 31: 339-369, 1991[ISI][Medline].

24.   McNicholas, CM, Wang W, Ho K, Hebert SC, and Giebisch G. Regulation of ROMK1 K+ channel activity involves phosphorylation processes. Proc Natl Acad Sci USA 91: 8077-8081, 1994[Abstract].

25.   Namba, T, Sugimoto Y, Negishi M, Irie A, Ushikubi F, Kakizuka A, Ito S, Ichikawa A, and Narumiya S. Alternative splicing of C-terminal tail of prostaglandin E receptor subtype EP3 determines G-protein specificity. Nature 365: 166-170, 1993[ISI][Medline].

26.   Palmer, LG, Choe H, and Frindt G. Is the secretory K channel in the rat CCT ROMK? Am J Physiol Renal Physiol 273: F404-F410, 1997[Abstract/Free Full Text].

27.   Peterson, LN, McKay AJ, and Borzecki JS. Endogenous prostaglandin E2 mediates inhibition of rat thick ascending limb Cl reabsorption in chronic hypercalcemia. J Clin Invest 91: 2399-2407, 1993[ISI][Medline].

28.   Reeves, WB, McDonald GA, Mehta P, and Andreoli TE. Activation of K+ channel in renal medullary vesicles by cAMP-dependent protein kinase. J Membr Biol 109: 65-72, 1989[ISI][Medline].

29.   Senba, S, Konishi K, Saruta T, Ozawa Y, Kato E, Amagasaki Y, and Nakata I. Hypokalemia and prostaglandin overproduction in Bartter's syndrome. Nephron 37: 257-263, 1984[ISI][Medline].

30.   Simon, DB, Karet FE, Rodriguez J, Hamdan JH, DiPietro A, Trachtman H, Sanjad SA, and Lifton RP. Genetic heterogeneity of Bartter's syndrome revealed by mutations in the K channel, ROMK. Nat Genet 14: 152-156, 1996[ISI][Medline].

31.   Stokes, JB. Effect of prostaglandin E on chloride transport across the rabbit thick ascending limb of Henle. J Clin Invest 64: 495-502, 1979[ISI][Medline].

32.   Taniguchi, J, and Guggino WB. Membrane stretch: a physiological stimulator of Ca2+-activated K+ channels in thick ascending limb. Am J Physiol Renal Fluid Electrolyte Physiol 257: F347-F352, 1989[Abstract/Free Full Text].

33.   Wang, WH. Two types of K+ channel in thick ascending limb of rat kidney. Am J Physiol Renal Fluid Electrolyte Physiol 267: F599-F605, 1994[Abstract/Free Full Text].

34.   Wang, WH, Hebert SC, and Giebisch G. Renal K channels: structure and function. Annu Rev Physiol 59: 413-436, 1997[ISI][Medline].

35.   Wang, WH, and Lu M. Effect of arachidonic acid on activity of the apical K channel in the thick ascending limb of the rat kidney. J Gen Physiol 106: 727-743, 1995[Abstract].

36.   Watabe, A, Sugimoto Y, Honda A, Irie A, Namba T, Negishi M, Ito S, Narumiya S, and Ichikawa A. Cloning and expression of cDNA for a mouse EP1 subtype of prostaglandin E receptor. J Biol Chem 268: 20175-20178, 1993[Abstract/Free Full Text].

37.   Xu, ZC, Yang Y, and Hebert SC. Phosphorylation of the ATP-sensitive, inwardly rectifying K+ Channel, ROMK, by cyclic AMP-dependent protein kinase. J Biol Chem 271: 9313-9319, 1996[Abstract/Free Full Text].


Am J Physiol Cell Physiol 278(5):C905-C913
0363-6143/00 $5.00 Copyright © 2000 the American Physiological Society