Circulating Levels Of Vasopressin Research Articles

Revisiting diuretic choice in chronic kidney disease.

Existing guidelines offer little direction about the use of thiazide and loop diuretics in patients with chronic kidney disease (CKD). This review summarizes recent studies impacting indications and safety considerations for these agents in patients with CKD. Chlorthalidone reduces blood pressure compared to placebo in patients with advanced CKD, challenging the belief that thiazide diuretics lose efficacy at lower glomerular filtration rates (GFR). Existing studies show no clear impact of thiazide or loop diuretic use on kidney or cardiovascular outcomes in patients with CKD. Sodium-glucose co-transporter type 2 (SGLT2) inhibitors have diuretic effects, but concomitant use of a diuretic does not diminish the preventive benefits of these agents against acute kidney injury (AKI). Despite theoretical concerns, thiazide diuretics likely do not worsen circulating vasopressin levels or cyst progression in polycystic kidney disease and may be useful for alleviating polyuria from tolvaptan. Diuretics cause multiple adverse effects, including electrolyte abnormalities, hemodynamic-mediated decrease in estimated GFR, and AKI. Recent evidence supports expanded indications for diuretics in patients with kidney disease, including chlorthalidone for hypertension in advanced CKD. Monitoring electrolytes and estimated GFR is critical to ensure patient safety when prescribing these agents for patients with CKD.

Vasopressin Receptor Expression Reveals Role of Endogenous Vasopressin in Ameliorating Acute Lung Injury in Pig Models of Pulmonary Hypertension

Acute lung injury (ALI) that evolves into acute respiratory distress syndrome (ARDS) is a critical care management challenge with 30–40% mortality. Despite the beneficial ability of pharmacological doses of vasopressin (VP)to elevate blood pressure without worsening pulmonary hypertension (PHN) in severe septic shock ARDS, circulating VP levels (not necessarily indicative of local tissue VP action) remain relatively low and it is unclear whether VP has a physiological role in maintaining pulmonary blood flow and oxygenation in ARDS. We hypothesized that if endogenous VP has an important role in ameliorating ALI, vascular VP receptor mRNA expression will increase within hours after initial ALI insult in order to replenish spent receptors that are degraded when activated by endogenous VP.Anesthetized Yorkshire cross pigs(n=36, 7.9 ± 0.2 kg body weight) were used to examine pulmonary hemodynamics in ALI without the confounding effects of systemic hypotension. PHN was induced by either disrupting the lung alveolar‐capillary interface with oleic acid (OA; 0.1 ml/kg iv, n=14) or by causing endothelial dysfunction with endotoxin (ETX; up to 35,000 units iv, n=16). ALI pigs had elevated pulmonary to systemic vascular resistance ratios (PVR/SVR: OA = 0.48 ± 0.07 and ETX = 0.48 ±0.04) compared to controls (0.20 ± 0.01, n=6). Mean arterial pressure was constant (60±4 mm Hg in all groups). ALI was managed with supplemental oxygen with or without mesenchymal stem cells to facilitate lung tissue repair. Within 2 hours after treatment, PVR/SVR began to return towards baseline in the ETX group but remained elevated in the OA group. This provided a range of pulmonary vasoconstriction states to compare against changes in plasma VP, pituitary VP content, and the mRNA expression of VP, V1aR, V1bR, and V2R receptors in pulmonary, carotid, renal, celiac, and mesenteric arteries, and aorta collected12 to 16 hours after initial ALI. Stepwise multiple regression revealed that PVR/SVR at 2–4 hours of established lung injury was positively correlated with V1aR expression in the aorta and mesenteric artery and V2R expression in the pulmonary and renal arteries, and negatively correlated with V2R expression in the aorta and celiac arteries (r2=0.88, p<0.0001). Results are consistent with endogenous VP responding to pulmonary hypertension in ALI via a pulmonary V2R mediated vasodilatory counter effect, in the face of V1aR mediated vasoconstriction in other vascular beds, to maintain pulmonary blood flow and improve blood oxygenation.Support or Funding InformationThis project was supported by the US Army Medical Command. The views expressed in this abstract are those of the authors and do not reflect the official policy or position of the Department of the Army, Department of Defense, or the U.S. GovernmentThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

CaSR signaling down-regulates AQP2 expression via a novel microRNA pathway in pendrin and NaCl cotransporter knockout mice.

High concentrations of urinary calcium counteract vasopressin action via the activation of the calcium-sensing receptor (CaSR) that is expressed in the luminal membrane of collecting duct cells, which impairs the trafficking of aquaporin-2 (AQP2). Pendrin/NaCl cotransporter double-knockout (dKO) mice display significant calcium wasting and develop severe volume depletion, despite increased circulating vasopressin levels. We hypothesized that the CaSR-mediated impairment of AQP2 expression/trafficking underlies vasopressin resistance in dKO mice. Compared with wild-type mice, in renal inner medulla, dKO mice had reduced total AQP2 sensitive to proteasome inhibitors, higher levels of AQP2-pS261, ubiquitinated AQP2, and p38-MAPK, an enzyme that is activated by CaSR signaling and known to phosphorylate AQP2 at Ser261. CaSR inhibition with the calcilytic NPS2143 reversed these effects, which indicates that CaSR mediates the up-regulation of AQP2-pS261, ubiquitination, and degradation. Of note, dKO mice demonstrated significantly higher AQP2-targeting miRNA-137 that was reduced upon CaSR inhibition, supporting a critical role for CaSR in the down-regulation of AQP2 expression. Our data indicate that CaSR signaling reduces AQP2 abundance both via AQP2-targeting miRNA-137 and the p38-MAPK/AQP2-pS261/ubiquitination/proteasomal axis. These effects may contribute to the reduced renal concentrating ability that has been observed in dKO mice and underscore a physiologic mechanism of the CaSR-dependent regulation of AQP2 abundance via a novel microRNA pathway.-Ranieri, M., Zahedi, K., Tamma, G., Centrone, M., Di Mise, A., Soleimani, M., Valenti, G. CaSR signaling down-regulates AQP2 expression via a novel microRNA pathway in pendrin and NaCl cotransporter knockout mice.

The Interaction of Vasopressin and Corticosteroids in Septic Shock

Vasopressin and corticosteroids are both commonly used adjunctive therapies in septic shock. Retrospective analyses have suggested that there may be an interaction between these drugs, with higher circulating vasopressin levels and improved outcomes in patients treated with both vasopressin and corticosteroids. We aimed to test for an interaction between vasopressin and corticosteroids in septic shock. Prospective open-label randomized controlled pilot trial. Four adult ICUs in London teaching hospitals. Sixty-one adult patients who had septic shock. Initial vasopressin IV infusion titrated up to 0.06 U/min and then IV hydrocortisone (50 mg 6 hourly) or placebo. Plasma vasopressin levels were measured at 6-12 and 24-36 hours after hydrocortisone/placebo administration. Thirty-one patients were allocated to vasopressin + hydrocortisone and 30 patients to vasopressin + placebo. The hydrocortisone group required a shorter duration of vasopressin therapy (3.1 d; 95% CI, 1.1-5.1; shorter in hydrocortisone group) and required a lower total dose of vasopressin (ratio, 0.47; 95% CI, 0.32-0.71) compared with the placebo group. Plasma vasopressin levels were not higher in the hydrocortisone group compared with the placebo group (64 pmol/L difference at 6- to 12-hour time point; 95% CI, -32 to 160 pmol/L). Early vasopressin use was well tolerated with only one serious adverse event possibly related to study drug administration reported. There were no differences in mortality rates (23% 28-day mortality in both groups) or organ failure assessments between the two treatment groups. Hydrocortisone spared vasopressin requirements, reduced duration, and reduced dose, when used together in the treatment of septic shock, but it did not alter plasma vasopressin levels. Further trials are needed to assess the clinical effectiveness of vasopressin as the initial vasopressor therapy with or without corticosteroids.

Pressor doses of vasopressin result in only transient elevations in plasma peptide levels

We recently reported that neuronostatin, a novel neuropeptide, biphasically increased mean arterial pressure, first through the activation of the sympathetic nervous system followed by the release of vasopressin. In those experiments, we found that centrally administered neuronostatin increased plasma vasopressin levels only 2–3 times greater than levels observed in saline-treated controls, and that the increase in mean arterial pressure (approximately 15mm Hg) could be blocked by pretreatment with a V1-vasopressin antagonist. Here we report the relationship between two to three fold elevations in plasma vasopressin levels and concomitant changes in mean arterial pressure in conscious, unrestrained male rats. We injected increasing doses of vasopressin (5, 20, and 100ng/kg, intra-arterially) and measured both changes in plasma vasopressin levels and the elevation in mean arterial pressure achieved. At 5-min post injection, plasma levels of vasopressin and mean arterial pressures were similar to those observed following central neuronostatin administration in our earlier study. Thus we conclude that small increases in circulating vasopressin levels can result in significant elevations in mean arterial pressure at least in the conscious rat.

Role of pendrin in iodide balance: going with the flow

Pendrin is expressed in the apical regions of type B and non-A, non-B intercalated cells, where it mediates Cl(-) absorption and HCO3(-) secretion through apical Cl(-)/HCO3(-) exchange. Since pendrin is a robust I(-) transporter, we asked whether pendrin is upregulated with dietary I(-) restriction and whether it modulates I(-) balance. Thus I(-) balance was determined in pendrin null and in wild-type mice. Pendrin abundance was evaluated with immunoblots, immunohistochemistry, and immunogold cytochemistry with morphometric analysis. While pendrin abundance was unchanged when dietary I(-) intake was varied over the physiological range, I(-) balance differed in pendrin null and in wild-type mice. Serum I(-) was lower, while I(-) excretion was higher in pendrin null relative to wild-type mice, consistent with a role of pendrin in renal I(-) absorption. Increased H2O intake enhanced differences between wild-type and pendrin null mice in I(-) balance, suggesting that H2O intake modulates pendrin abundance. Raising water intake from approximately 4 to approximately 11 ml/day increased the ratio of B cell apical plasma membrane to cytoplasm pendrin label by 75%, although circulating renin, aldosterone, and serum osmolality were unchanged. Further studies asked whether H2O intake modulates pendrin through the action of AVP. We observed that H2O intake modulated pendrin abundance even when circulating vasopressin levels were clamped. We conclude that H2O intake modulates pendrin abundance, although not likely through a direct, type 2 vasopressin receptor-dependent mechanism. As water intake rises, pendrin becomes increasingly critical in the maintenance of Cl(-) and I(-) balance.

Stimulation of adenosine A1 receptors in the nucleus of the solitary tract (NTS) triggers the release of vasopressin into the circulation

Our previous study showed that stimulation of NTS A1 adenosine receptors exerts counteracting effects on the iliac vascular bed: activation of the adrenal medulla and β‐adrenergic vasodilation vs. vasoconstriction mediated by neural and unknown humoral factors (Am J Physiol 289:H2536‐H2542, 2005). The follow up study identified vasopressin as a potential humoral factor involved: systemic blockade of V1 vasopressin receptors abolished/reversed iliac vasoconstriction (FASEB 20: A363, 2006). Stimulation of NTS A1 adenosine receptors attenuates the baroreflex (FASEB 21: A466, 2007), thus may increase circulating vasopressin. Therefore in the present study we compared circulating vasopressin levels before and after microinjections into the NTS of selective A1 adenosine receptor agonist N6cyclopentyladenosine (CPA, 330 pmol/50nl) or vehicle (artificial cerebrospinal fluid, ACF, 50 nl), using the same experimental model as previously (chloralose/urethane anesthetized rats). CPA increased circulating vasopressin over 4‐fold (117.0±19.0 vs. 26.4±10.4 pg/ml, P=0.0006, n=8) whereas ACF did not increase vasopressin level significantly (50.9±11.3 vs. 25.0±7.2 pg/ml, P=0.083, n=5). We conclude that stimulation of NTS A1 adenosine receptors increases circulating vasopressin probably via inhibition of NTS baroreflex mechanisms. NIH HL‐67814

Renal function in transgenic rats expressing an angiotensin-(1–7)-producing fusion protein

Transgenic rats [TGR(A1–7)3292] present a chronic 2.5-fold increase in plasma Angiotensin-(1–7) [Ang-(1–7)] concentration. In the present study, we investigated the effects of this chronic elevation on renal function, vasopressin levels, kidney morphology, expression of Ang-(1–7) and vasopressin receptors in TGR(A1–7)3292. Urine volume and water intake were measured for 24 h. At the end of this period, plasma and urine samples were collected to evaluate renal function parameters and circulating vasopressin levels. Expression of renal V 2 receptors and Mas was assessed by ribonuclease protection assay. Renal slices were processed for histological analysis. The urine flow of TGR(A1–7)3292 was significantly lower in comparison with Sprague–Dawley rats. The reduced urine volume of TGR(A1–7)3292 was accompanied by a significant increase in urinary osmolality and decrease free water clearance. Glomerular filtration rate, urinary sodium and potassium excretion were similar in both strains. No significant changes were observed in vasopressin levels as well as in V 2 receptor and Mas mRNA expression in renal tissue. No changes in kidney structure of TGR(A1–7)3292 were detected. These data suggest that changes in circulating renin–angiotensin system produced by chronic increase of Ang-(1–7) levels can lead to adjustments in the water balance that are independent of vasopressin release and V 2 receptor expression.

Serial circulating vasopressin levels in children with septic shock*

Septic shock is an important cause of death in pediatric intensive care units. Initial evaluations have shown that vasopressin may have a role in catecholamine refractory shock in adults. It is important to determine whether children with septic shock have deficiency of vasopressin. This will help in defining the role of vasopressin in septic shock. Prospective cohort study. Pediatric intensive care unit of a tertiary care hospital in north India. Patients were children with septic shock, and controls were children with sepsis but no shock. Vasopressin levels in plasma were determined by enzyme-linked immunosorbent assay for children with septic shock at diagnosis (baseline) and thereafter at 24, 48, and 96 hrs to determine the time trends. The baseline vasopressin values for children with septic shock were compared with those for children without shock. The median (95% confidence interval) vasopressin level at baseline in children with septic shock was 116 (63.3-130.7) pg/mL, and in children with sepsis but no shock it was 106 (61.7-131.77) pg/mL. The median value for survivors was 76 (44.6-130.9) pg/mL, and for nonsurvivors, 118 (81.7-259) pg/mL (p = .16). The serial values also did not show any significant changes; the values at 24 hrs (n = 17), 48 hrs (n = 16), and 96 hrs (n = 15) were 105 (76.1-125.9), 105 (41.4-155.5), and 109.5 (54.9-154.8) pg/mL, respectively. The results of our study suggest that vasopressin levels are elevated in children with septic shock and that serial values up to 96 hrs do not show any decline.

Serial circulating vasopressin levels in children with septic shock

Serial circulating vasopressin levels in children with septic shock

Serial circulating vasopressin levels in children with septic shock

Serial circulating vasopressin levels in children with septic shock

P2Y2 receptor-mediated release of prostaglandin E2by IMCD is altered in hydrated and dehydrated rats: relevance to AVP-independent regulation of IMCD function

Circulating vasopressin levels change in hydrated and dehydrated conditions and thus control osmotic water permeability (P(f)) of the inner medullary collecting duct (IMCD). Prostaglandin E2 (PGE2) antagonizes vasopressin-induced P(f) of IMCD. Previously, we showed that activation of P2Y2 receptor (P2Y2-R) in IMCD results in production and release of PGE2, and P2Y2-R mRNA and protein are significantly elevated in inner medullas of hydrated rats compared with dehydrated rats. Therefore, we examined whether the altered expression of P2Y2-R in hydrated and dehydrated states is associated with corresponding changes in P2Y2-R-mediated PGE2 release by the IMCD. Rats were hydrated by providing sucrose water as the sole drinking fluid or dehydrated by water deprivation for 2 days. This resulted in high output-low osmolality and low output-high osmolality urines in hydrated and dehydrated rats, respectively. In hydrated rats, there was a significant increase in tubular fluid PGE2, measured indirectly by assessing the urinary PGE2 metabolite. Stimulation of freshly isolated IMCD preparations in vitro with P2Y2-R agonist (ATPgammaS) showed a marked increase in the release of PGE2 in hydrated rats compared with normal rats. These responses were blunted in the IMCD prepared from dehydrated rats. The P2Y2-R-mediated PGE2 release in the IMCD of hydrated rats was mediated largely by cyclooxygenase (COX)-1 as COX-1-specific inhibitor valeroyl salicylate completely blocked the release. The COX-2-specific inhibitor N5398 had only a modest and insignificant inhibitory effect. In conclusion, the increased sensitivity of purinergic-prostanoid interaction seen in the IMCD of hydrated rats may represent a novel vasopressin-independent regulatory mechanism of IMCD function.

Downregulation of renal AQP2 water channel and NKCC2 in mice lacking the apical Na+-H+ exchanger NHE3.

The apical Na+-H+ exchanger NHE3 plays an important role in fluid reabsorption in the proximal tubule. However, whether its deletion alters the salt and water transport in the distal nephron remains unknown. To answer these questions, wild-type (Nhe3+/+) and NHE3 null mice (Nhe3-/-) were placed in metabolic cages and their water balance and urine osmolality were examined. Nhe3-/- mice demonstrated a significant polydipsia (P < 0.03) and polyuria (P < 0.04), with a lower urine osmolality (P < 0.003) as compared to Nhe3+/+ mice. Northern hybridization and immunoblotting studies indicated that the mRNA expression and protein abundance of the collecting duct (CD) water channel AQP2 decreased by 52 % (P < 0.0003) and 73 % (P < 0.003) in the cortex, and by 53 % and 54 % (P < 0.002) in the inner medulla (IM) of Nhe3-/- vs. Nhe3+/+ mice. The expression of AQP2 in the outer medulla (OM) remained unchanged. Further, the mRNA expression and protein abundance of the medullary thick ascending limb (mTAL) apical Na+-K+-2Cl- cotransporter (NKCC2) decreased by 52 % (P < 0.02) and 44 % (P < 0.01), respectively, in the OM of Nhe3-/- vs. Nhe3+/+ mice. The circulating plasma levels of vasopressin as well as the mRNA expression of vasopressin prohormone were significantly increased in Nhe3-/- vs. Nhe3+/+ mice (P < 0.05). Studies in mice treated with acetazolamide indicated that increased bicarbonate and fluid delivery to distal nephron did not alter the expression of NKCC2 in mTAL and decreased AQP2 protein only in OM but not in the cortex or IM. In conclusion, mice lacking the apical NHE3 have impairment in their water balance and urine osmolality, which correlates with the downregulation of AQP2 expression. These defects occur despite increased circulating levels of vasopressin. We propose that an ADH-independent mechanism is responsible for the downregulation of AQP2 and the resulting polyuria in NHE3 null mice.

Circulating vasopressin levels in septic shock.

To assess the frequency of vasopressin deficiency in septic shock. Prospective cohort study. Intensive care unit at Raymond Poincaré University Hospital. A cohort of 44 patients who met the usual criteria for septic shock for < 7 days. A second cohort of 18 septic shock patients were enrolled within the first 8 hrs of disease onset. None. General demographics, severity scores, vital signs, standard biochemical data, and circulating vasopressin levels were systematically obtained at baseline in the two cohorts. Vasopressin deficiency was defined by a normal plasma vasopressin level in the presence of a systolic blood pressure of <100 mm Hg or in the presence of hypernatremia. Baroreflex sensitivity was systematically evaluated in patients of the first cohort when vasopressin deficiency was noted. In the second cohort of patients, plasma levels of vasopressin were obtained at baseline, 6, 24, 48, and 96 hrs after shock onset. In the first population, plasma vasopressin levels were inversely correlated to the delay from shock onset. Fourteen patients had relative vasopressin deficiency: 12 patients had systolic blood pressure <100 mm Hg, with impaired baroreflex sensitivity in four, and three patients had hypernatremia. In the second population, only two patients had relative vasopressin deficiency. The plasma levels of vasopressin significantly decreased over time (p < 10-3). Plasma vasopressin levels are almost always increased at the initial phase of septic shock and decrease afterward. Relative vasopressin deficiency is seen in approximately one-third of late septic shock patients.

Mechanisms involved in the pressor response to noradrenaline injection into the cingulate cortex of unanesthetized rats

The cingulate cortex (CC) is involved in cardiovascular modulation. CC electrical or chemical stimulation may evoke either pressor or depressor responses, depending on the stimulated site and experimental conditions such as anesthesia. Noradrenaline (NA) is involved in cardiovascular regulation and it is present throughout the cortex. However, there is no report on the cardiovascular effects of intracortical injections of NA. We attempted to verify the effect of NA injection into the CC and to identify possible receptor and peripheral mechanisms involved. NA injection caused pressor responses accompanied by bradycardia, in unanesthetized rats. These responses were markedly reduced under urethane anesthesia. The pressor response was blocked by intracortical pretreatment with phenoxybenzamine or the selective α 1-antagonist WB4101, and it was not affected by pretreatment with the selective α 2-antagonist RX821002, suggesting that α 1-adrenoceptors mediate the response. The pressor response was potentiated by pretreatment with the ganglion blocker mecamylamine and it was abolished by pretreatment with the vasopressin antagonist, dTyr(CH 2) 5(Me)AVP or by hypophysectomy. Circulating vasopressin levels were increased after NA injection into the CC. The present results indicate that the pressor response to local injection of NA within the CC is independent of sympathetic nerve activation and is mediated by vasopressin release.

Nitric oxide, interleukin and prostaglandin interactions affecting the magnocellular system

Magnocellular neurons are innervated by an excitatory histaminergic pathway. They also express neuronal NO synthase, interleukin-1β (IL-1β) and cyclo-oxygenase (COX). In normally hydrated rats when NO synthase activity is inhibited with N G -nitro- l-arginine methyl ester ( l-NAME), administered intracerebroventricularly (i.c.v.), OT concentration in plasma increases. In the present study, the increase in hormone after l-NAME is attenuated by indomethacin, an inhibitor of COX, as well as by antagonists of histamine receptors at H 1 (pyrilamine) and H 2 (cimetidine) subtypes injected i.c.v. Moreover, enhanced OT secretion induced by centrally administered IL-1β, but not naloxone (opiate receptor antagonist), is prevented by indomethacin. PGE 2 and PGD 2 (i.c.v.) stimulate OT release, but only PGD 2 affects circulating vasopressin levels. Thus, NO inhibits release of OT stimulated by: (1) a COX-dependent mechanism, i.e. NO→−(COX→+PG→+OT release); (2) histamine, i.e. NO→−(histamine→H 1 and H 2 receptors→+OT release); and possibly (3) IL-1β, i.e. NO→−(IL-1β→+COX→+PG→+OT release). These interactions of NO, cytokine and histamine may be important for management of stress-induced activation of neuroendocrine systems.

Chapter 6 Regulation of renal salt and water transporters during vasopressin escape

Hyponatremia, defined as a serum sodium < 135 mmol/l, is one of the most commonly encountered and serious electrolyte disorders of clinical medicine. The predominant cause of hyponatremia is an inappropriate elevation of circulating vasopressin levels relative to serum osmolality or the 'syndrome of inappropriate antidiuretic hormone secretion' (SIADH). Fortunately, the degree of the hyponatremia is limited by a process that counters the water-retaining action of vasopressin, namely 'vasopressin escape'. Vasopressin escape is characterized by a sudden increase in urine volume with a decrease in urine osmolality independent of circulating vasopressin levels. Until recently, little was known about the molecular mechanisms underlying escape. In the 1980s, we developed an animal model for vasopressin escape in which male Sprague-Dawley rats were infused with dDAVP, a V2-receptor-selective agonist of vasopressin, while being fed a liquid diet. Rats drank a lot of water in order to get the calories they desired. Using this model, we demonstrated that the onset of vasopressin escape (increased urine volume coupled to decreased urine osmolality) coincided temporally with a marked decrease in renal aquaporin-2 (water channel) protein and mRNA expression in renal collecting ducts. This protein reduction was reversible and correlated to decreased water permeability of the collecting duct. Studies examining the mechanisms underlying AQP2 decrease revealed a decrease in V2-receptor mRNA expression and binding, as well as a decrease in cyclic AMP production in response to acute-dDAVP challenge in collecting duct suspensions from these escape animals. Additional studies showed an increase in sodium transporters of the distal tubule. These changes, hypothetically, might help to attenuate the hyponatremia. Future studies are needed to fully elucidate systemic, intra-organ, and cellular signaling responsible for the physiological phenomenon of vasopressin escape.

Targeted proteomics in the kidney using ensembles of antibodies.

Building on extensive physiological characterization of sodium transport mechanisms along the renal tubule over the past 30 years, complementary DNAs for almost all of the major transporters and channels responsible for renal tubular sodium reabsorption have been cloned over the past 10 years. The consequence is the generation of a broad range of cDNA and antibody probes which can be used to investigate physiological mechanisms on a molecular level. An ensemble of such probes can be exploited for comprehensive analysis of integrative physiological processes, approaches which are referred to as 'physiological genomics' or 'physiological proteomics'. In this review, we describe a targeted proteomic approach to comprehensive analysis of sodium transporter and water channel protein abundance along the renal tubule using an ensemble of rabbit polyclonal antibodies directed to the major sodium transporters and water channels expressed in each renal tubule segment. We discuss preparation and characterization of the antibodies, strategies for quantification of transporter protein abundance, and provide examples of the application of antibody-based targeted proteomics analysis of kidney tissue, revealing the effects of elevations of circulating aldosterone levels and circulating vasopressin levels on sodium transporter, sodium channel, and water channel abundance in kidney.

A functional interaction between the mesolimbic dopamine system and vasopressin release in the regulation of blood pressure in conscious rats

The aim of the present study was to further characterize the involvement of the mesolimbic dopamine system in central blood pressure regulation, with particular emphasis on the interaction of this system with the effects of circulating vasopressin. In conscious rats we stimulated the release of endogenous dopamine from mesolimbic/mesocortical terminals by administration of the substance P analogue DiMe-C7 ([pGlu 5, MePhe 8, Sar 9]-Substance P 5–11; 10 nmol) into the ventral tegmental area. Chemical stimulation of the ventral tegmental area resulted in a significant increase in blood pressure and heart rate. These effects were prevented by either bilateral electrolytic lesions of the hypothalamic supraoptic nucleus or by systemic pretreatment with the dopamine D 2 receptor antagonist raclopride (0.5 mg/kg). Stimulation of the ventral tegmental area also produced a marked increase in the expression of the proto-oncogene c-fos in the supraoptic nucleus and a significant increase in plasma vasopressin levels, suggesting activation of vasopressinergic neurons in this nucleus. However, this effect of stimulation of the ventral tegmental area was not significantly inhibited by pretreatment with raclopride. We suggest that the effects on blood pressure and heart rate of stimulation of the ventral midbrain by micro-injection of DiMe-C7 are the result of combined activation of both dopaminergic and non-dopaminergic cell bodies in this region. Stimulation of non-dopaminergic cells in the ventral midbrain may induce a moderate increase in plasma vasopressin levels by activation of the supraoptic nucleus. An additional stimulation of dopaminergic cells in the ventral midbrain allows the increase in circulating vasopressin levels to become manifest as a pressor response, possibly by inhibition of vasopressin-induced facilitation of baroreflex responses.

Redistribution of aquaporin-2 water channels induced by vasopressin in rat kidney inner medullary collecting duct.

Aquaporin-2 (AQP2) is the predominant vasopressin-regulated water channel of the renal collecting duct. We tested whether vasopressin induces translocation of AQP2 from intracellular vesicles into the apical plasma membrane. AQP2 was quantitated in plasma membrane and intracellular vesicle fractions prepared from the inner medulla of one kidney from each rat before or 20 min after intravenous 1-desamino-8-D-arginine vasopressin (DDAVP) treatment, using immunoblotting and densitometry. Contralateral kidneys were prepared for immunofluorescence and immunoelectron microscopy. Immunoblotting revealed that, compared with untreated controls, DDAVP treatment significantly increased the fraction of AQP2 protein associated with the plasma membrane fraction relative to intracellular vesicles. This increase averaged 2.0-fold in untreated rats and 2.9-fold in rats water loaded for 12 h. Water loading, presumably by suppressing circulating vasopressin levels, decreased the fraction of AQP2 associated with the plasma membrane by 55%, suggesting retrieval of AQP2 from the plasma membrane. In rats sequentially thirsted for 48 h to increase expression and then water loaded for 72 h to minimize plasma membrane labeling, DDAVP caused a 12-fold increase in the plasma membrane to intracellular vesicle labeling ratio. The accentuation of the DDAVP response seen after water loading is consistent with the observed increase in the fraction of AQP2 in the intracellular pool available for insertion. Immunofluorescence confirmed a marked DDAVP-induced redistribution of AQP2 from intracellular to plasma membrane domains. Furthermore, quantitative immunoelectron microscopy demonstrated a 3.4-fold increase in apical plasma membrane to intracellular vesicle labeling ratio. These results provide a direct in vivo demonstration of vasopressin-induced translocation of AQP2 into the apical plasma membrane.