Vasopressin V2 receptor mediated Ca2+ transients in the rat inner medullary collecting duct are dependent on phospholipase C and extracellular Ca2+

Phosphorylation and lysine (K)-acetylation are dynamic posttranslational modifications of proteins. Previous proteomic studies have identified over 170,000 phosphorylation sites and 15,000 K-acetylation sites in mammals. We recently reported that the inner medullary collecting duct (IMCD), which functions in the regulation of water-reabsorption, via the actions of vasopressin, expresses many of the enzymes that can modulated K-acetylation. The purpose of this study was to determine the K-acetylated or phosphorylated proteins expressed in IMCD cells. Second we questioned whether vasopressin V2 receptor activation significantly affects the IMCD acetylome or phosphoproteome? K-acetylated or serine-, threonine-, or tyrosine-phosphorylated peptides were identified from native rat IMCDs by proteomic analysis with four different enzymes (trypsin, chymotrypsin, ASP-N, or Glu-C) to generate a high-resolution proteome. K-acetylation was identified in 431 unique proteins, and 64% of the K-acetylated sites were novel. The acetylated proteins were expressed in all compartments of the cell and were enriched in pathways including glycolysis and vasopressin-regulated water reabsorption. In the vasopressin-regulated water reabsorption pathway, eight proteins were acetylated, including the novel identification of the basolateral water channel, AQP3, acetylated at K282; 215 proteins were phosphorylated in this IMCD cohort, including AQP2 peptides that were phosphorylated at four serines: 256, 261, 264, and 269. Acute dDAVP did not significantly affect the IMCD acetylome; however, it did significantly affect previously known vasopressin-regulated phosphorylation sites. In conclusion, presence of K-acetylated proteins involved in metabolism, ion, and water transport in the IMCD points to multiple roles of K-acetylation beyond its canonical role in transcriptional regulation.

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Vasopressin and oxytocin receptors coupled to Ca2+ mobilization in rat inner medullary collecting duct

AJP Renal Physiology ◽

10.1152/ajprenal.1993.265.1.f15 ◽

1993 ◽

Vol 265 (1) ◽

pp. F15-F25 ◽

Cited By ~ 11

Author(s):

Y. Maeda ◽

J. S. Han ◽

C. C. Gibson ◽

M. A. Knepper

Keyword(s):

Arginine Vasopressin ◽

Receptor Agonist ◽

Collecting Duct ◽

Oxytocin Receptor ◽

Receptor Subtype ◽

Vasopressin Receptor ◽

Inner Medullary Collecting Duct ◽

V1b Receptor ◽

V2 Receptor ◽

Medullary Collecting Duct

In renal collecting duct epithelial cells, arginine vasopressin (AVP) at greater than nanomolar concentrations has been reported to transiently increase intracellular free calcium ([Ca2+]i) in a manner consistent with activation of the phosphoinositide pathway. To investigate whether any of the known neurohypophysial hormone subtypes are involved, we measured [Ca2+]i in microdissected rat terminal inner medullary collecting duct (IMCD) using fura-2. To allow quantitative comparisons of the response under different conditions, we determined the areas under the response curves (in nM.min) over 1.5 min using numerical integration. AVP, the V1b-receptor agonist [deamino1,D-3-(pyridyl)Ala2,Arg8]vasopressin, the V2-receptor agonist 1-desamino-8-D-arginine vasopressin, oxytocin, and the selective oxytocin-receptor agonist [Thr4,Gly7]oxytocin (TG-OXT), each at 10 nM, significantly increased [Ca2+]i (69.52 +/- 10.25, 27.0 +/- 11.7, 24.33 +/- 5.83, 14.75 +/- 2.81, and 14.57 +/- 3.50 nM.min, respectively). In contrast, a V1a-selective agonist ([Phe2,Ile3,Orn8]vasopressin) did not increase [Ca2+]i (0.43 +/- 2.36 nM.min). In desensitization studies, challenge with 10 nM AVP or TG-OXT completely prevented a rise in [Ca2+]i in response to immediate rechallenge with the same agent, but not the other, demonstrating homologous desensitization. The lack of cross-desensitization implies that at least two receptors are present that can trigger a rise in [Ca2+]i in response to neurohypophysial hormones. Antagonists for oxytocin ([des-glycinamide9,d(CH2)5(1),O-Me-Tyr2,Thr4,Orn8]vaso tocin), V2 ([d(CH2)5(1),D-Ile2,Ile4,Arg8]vasopressin), and V1a ([d(CH2)5(1),O-Me-Tyr2,Arg8]vasopressin) receptors partially inhibited the [Ca2+]i response induced by 10 nM AVP (89.5, 81.6, and 51.4% inhibition, respectively). These data are consistent with the view that both an oxytocin receptor and a vasopressin receptor are coupled to a [Ca2+]i mobilization response in rat terminal IMCD. This vasopressin receptor is distinct from both the V1a receptor and the V2 receptor and may be either the V1b receptor or a novel vasopressin receptor subtype.

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Evidence for dual signaling pathways for V2 vasopressin receptor in rat inner medullary collecting duct

AJP Renal Physiology ◽

10.1152/ajprenal.1996.270.4.f623 ◽

1996 ◽

Vol 270 (4) ◽

pp. F623-F633 ◽

Cited By ~ 34

Author(s):

C. A. Ecelbarger ◽

C. L. Chou ◽

S. J. Lolait ◽

M. A. Knepper ◽

S. R. DiGiovanni

Keyword(s):

Arginine Vasopressin ◽

Receptor Agonist ◽

Collecting Duct ◽

Calcium Mobilization ◽

Mrna Levels ◽

Inner Medullary Collecting Duct ◽

Receptor Mrna ◽

V1a Receptor ◽

V2 Receptor ◽

Medullary Collecting Duct

Previous studies have demonstrated that both the V2-receptor agonist, 1-desamino-8-D-arginine vasopressin (dDAVP), and the V1a-receptor agonist, [Phe2, Orn8]vasotocin (PO-VT), increase intracellular calcium concentration ([Ca2+]i) in the rat inner medullary collecting duct (IMCD). The present studies were done to clarify the receptor subtype(s) responsible for calcium mobilization. Measurements of [Ca2+]i, using fura 2 in microdissected IMCD segments, confirmed that arginine vasopressin (AVP), dDAVP, and PO-VT stimulate an increase in [Ca2+]i and that the response to all three agents could be blocked by the specific V2-receptor antagonist, [d(CH2)5(1),D-Ile2, Ile4, Arg8]vasopressin (II-VP). These results would suggest that all three agents acted through the V2 receptor. Furthermore, we showed that PO-VT increased cAMP production in IMCD suspensions and water permeability in isolated perfused tubules. These responses were also blocked by II-VP, indicating that PO-VT is also a V2 agonist in the IMCD. Finally, we utilized the quantitative reverse transcription-polymerase chain reaction technique of Wiesner (Nucleic Acids Res. 20: 5863-5864, 1992) to evaluate V1a and V2 mRNA levels in rat collecting duct. In terminal IMCD, we estimated > 30 copies/cell for V2 receptor mRNA but less than 1 copy/cell of V1a receptor mRNA, thus there is littler or no V1a mRNA expression in the terminal IMCD. These results suggest that calcium mobilization in response to vasopressin analogues is associated with the V2 receptor and that the V2 receptor is linked to both adenylyl cyclase and calcium mobilization in the rat IMCD.

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The Role of Vasopressin V2 Receptor in Drug-Induced Hyponatremia

Frontiers in Physiology ◽

10.3389/fphys.2021.797039 ◽

2021 ◽

Vol 12 ◽

Author(s):

Sua Kim ◽

Chor Ho Jo ◽

Gheun-Ho Kim

Keyword(s):

Collecting Duct ◽

Animal Experiments ◽

Urine Concentration ◽

Cytotoxic Agents ◽

Drug Induced ◽

Vasopressin V2 Receptor ◽

Posterior Pituitary Gland ◽

V2 Receptor ◽

Collecting Duct Cells ◽

Medullary Collecting Duct

Hyponatremia is frequently encountered in clinical practice and usually induced by renal water retention. Many medications are considered to be among the various causes of hyponatremia, because they either stimulate the release of arginine vasopressin (AVP) or potentiate its action in the kidney. Antidepressants, anticonvulsants, antipsychotics, diuretics, and cytotoxic agents are the major causes of drug-induced hyponatremia. However, studies addressing the potential of these drugs to increase AVP release from the posterior pituitary gland or enhance urine concentration through intrarenal mechanisms are lacking. We previously showed that in the absence of AVP, sertraline, carbamazepine, haloperidol, and cyclophosphamide each increased vasopressin V2 receptor (V2R) mRNA and aquaporin-2 (AQP2) protein and mRNA expression in primary cultured inner medullary collecting duct cells. The upregulation of AQP2 was blocked by the V2R antagonist tolvaptan or protein kinase A (PKA) inhibitors. These findings led us to conclude that the nephrogenic syndrome of inappropriate antidiuresis (NSIAD) is the main mechanism of drug-induced hyponatremia. Previous studies have also shown that the V2R has a role in chlorpropamide-induced hyponatremia. Several other agents, including metformin and statins, have been found to induce antidiuresis and AQP2 upregulation through various V2R-independent pathways in animal experiments but are not associated with hyponatremia despite being frequently used clinically. In brief, drug-induced hyponatremia can be largely explained by AQP2 upregulation from V2R-cAMP-PKA signaling in the absence of AVP stimulation. This paper reviews the central and nephrogenic mechanisms of drug-induced hyponatremia and discusses the importance of the canonical pathway of AQP2 upregulation in drug-induced NSIAD.

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Na+/H+ exchanger isoforms NHE-2 and NHE-1 in inner medullary collecting duct cells. Expression, functional localization, and differential regulation.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(18)46882-2 ◽

1994 ◽

Vol 269 (45) ◽

pp. 27973-27978

Author(s):

M Soleimani ◽

G Singh ◽

G L Bizal ◽

S R Gullans ◽

J A McAteer

Keyword(s):

Collecting Duct ◽

Differential Regulation ◽

Inner Medullary Collecting Duct ◽

Duct Cells ◽

Functional Localization ◽

Collecting Duct Cells ◽

Medullary Collecting Duct

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Water permeability of apical and basolateral cell membranes of rat inner medullary collecting duct

AJP Renal Physiology ◽

10.1152/ajprenal.1990.259.6.f986 ◽

1990 ◽

Vol 259 (6) ◽

pp. F986-F999 ◽

Cited By ~ 5

Author(s):

B. Flamion ◽

K. R. Spring

Keyword(s):

Water Flow ◽

Water Permeability ◽

Cell Membranes ◽

Apical Membrane ◽

Collecting Duct ◽

Basolateral Membrane ◽

Volume Regulatory Decrease ◽

Water Permeation ◽

Inner Medullary Collecting Duct ◽

Medullary Collecting Duct

To quantify the pathways for water permeation through the kidney medulla, knowledge of the water permeability (Posmol) of individual cell membranes in inner medullary collecting duct (IMCD) is required. Therefore IMCD segments from the inner two thirds of inner medulla of Sprague-Dawley rats were perfused in vitro using a setup devised for rapid bath and luminal fluid exchanges (half time, t1/2, of 55 and 41 ms). Differential interference contrast microscopy, coupled to video recording, was used to measure volume and approximate surface areas of single cells. Volume and volume-to-surface area ratio of IMCD cells were strongly correlated with their position along the inner medullary axis. Transmembrane water flow (Jv) was measured in response to a variety of osmotic gradients (delta II) presented on either basolateral or luminal side of the cells. The linear relation between Jv and delta II yielded the cell membrane Posmol, which was then corrected for membrane infoldings. Basolateral membrane Posmol was 126 +/- 3 microns/s. Apical membrane Posmol rose from a basal value of 26 +/- 3 microns/s to 99 +/- 5 microns/s in presence of antidiuretic hormone (ADH). Because of amplification of basolateral membrane, the ADH-stimulated apical membrane remained rate-limiting for transcellular osmotic water flow, and the IMCD cell did not swell significantly. Calculated transcellular Posmol, expressed in terms of smooth luminal surface, was 64 microns/s without ADH and 207 microns/s with ADH. IMCD cells in anisosmotic media displayed almost complete volume regulatory decrease but only partial volume regulatory increase.

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