Marcelo R. Choi

Signaling pathways involved in atrial natriuretic factor and dopamine regulation of renal Na+, K+ -ATPase activity.

Dopamine (DA) and atrial natriuretic factor (ANF) share a number of physiological effects. We hypothesized that ANF and the renal dopaminergic system could interact and enhance the natriuretic and diuretic effects of the peptide. We have previously reported that the ANF-stimulated DA uptake in renal tubular cells is mediated by the natriuretic peptide type-A receptor (NPR-A). Our aim was to investigate the signaling pathways that mediate ANF effects on renal 3H-DA uptake. Methylene blue (10 microM), an unspecific inhibitor of guanylate cyclase (GC), blunted ANF elicited increase of DA uptake. ODQ (10 microM) a specific inhibitor of soluble GC, did not modify DA uptake and did not reverse ANF-induced increase of DA uptake; then the participation of nitric oxide-dependent pathways must be discarded. The second messenger was the cGMP since the analogous 125 microM 8-Br-cGMP mimicked ANF effects. The specific inhibitor of the protein kinase G (PKG), KT 5823 (1 microM) blocked ANF effects indicating that PKG is involved. We examined if ANF effects on DA uptake were able to modify Na+, K+ -adenosine triphosphatase (Na+, K+ -ATPase) activity. The experiments were designed by means of inhibition of renal DA synthesis by carbidopa and neuronal DA uptake blocked by nomifensine. In these conditions renal Na+, K+ -ATPase activity was increased, in agreement with the decrease of DA availability. When in similar conditions, exogenous DA was added to the incubation medium, the activity of the enzyme tended to decrease, following to the restored availability of DA. The addition of ANF alone had similar effects to the addition of DA on the sodium pump, but when both were added together, the activity of Na(+), K(+)-ATPase was decreased. Moreover, the extraneuronal uptake blocker, hydrocortisone, inhibited the latter effect. In conclusion, ANF stimulates extraneuronal DA uptake in external cortex tissues by activation of NPR-A receptors coupled to GC and it signals through cGMP as second messenger and PKG. Dopamine and ANF may achieve their effects through a common pathway that involves reversible deactivation of renal tubular Na+, K+ -ATPase activity. This mechanism demonstrates a DA-ANF relationship involved in the modulation of both decreased sodium reabsorption and increased natriuresis.

Urodilatin and dopamine: A new interaction in the kidney

Since renal natriuretic peptide urodilatin (URO) exerts similar natriuretic and diuretic actions to those of atrial natriuretic factor (ANF), we hypothesized that URO regulates renal dopamine (DA) availability, contributing to Na(+), K(+)-ATPase inhibition. URO (1-100 nM) increased (3)H-DA uptake in outer and juxtamedullar renal cortex and medulla slices from Sprague Dawley rats. Hydrocortisone blocked URO-stimulated DA uptake, demonstrating that DA uptake was extraneuronal. The natriuretic peptide receptor type A antagonist anantin blocked URO-dependent increase of (3)H-DA uptake, while the natriuretic peptide receptor type C agonist ANF 4-23-amide did not modify URO effect on DA uptake, suggesting that only natriuretic receptors type A are involved. Co-incubation of URO and ANF did not show additive effects on DA uptake. To test whether URO effect involves changes in Na(+), K(+)-ATPase activity we performed experiments in renal cortex samples of rats with DA synthesis and neuronal uptake inhibited by carbidopa and nomifensine, respectively. When endogenous DA synthesis was inhibited, URO or DA decreased Na(+), K(+)-ATPase activity. URO and DA added together, further decreased Na(+), K(+)-ATPase activity showing an additive effect on the sodium pump. Moreover, hydrocortisone reversed URO-DA over-inhibition of the enzyme, confirming that this inhibition is closely related to URO-stimulation on renal DA uptake. URO and DA could act via a common intracellular pathway to decrease sodium and water tubular reabsorption, contributing to its natriuretic and diuretic effects.

Renal dopaminergic system: Pathophysiological implications and clinical perspectives

Fluid homeostasis, blood pressure and redox balance in the kidney are regulated by an intricate interaction between local and systemic anti-natriuretic and natriuretic systems. Intrarenal dopamine plays a central role on this interactive network. By activating specific receptors, dopamine promotes sodium excretion and stimulates anti-oxidant and anti-inflammatory pathways. Different pathological scenarios where renal sodium excretion is dysregulated, as in nephrotic syndrome, hypertension and renal inflammation, can be associated with impaired action of renal dopamine including alteration in biosynthesis, dopamine receptor expression and signal transduction. Given its properties on the regulation of renal blood flow and sodium excretion, exogenous dopamine has been postulated as a potential therapeutic strategy to prevent renal failure in critically ill patients. The aim of this review is to update and discuss on the most recent findings about renal dopaminergic system and its role in several diseases involving the kidneys and the potential use of dopamine as a nephroprotective agent.

Angiotensin II Regulation of Renal Dopamine Uptake and Na+,K+-ATPase Activity

Background/Aims: Angiotensin II (ANG II) decreases dopamine (DA) uptake in renal cortex activating AT₁ receptors. We investigated the signaling pathways that mediate this action and the incidence of DA-ANG II interaction on renal Na⁺,K⁺-ATPase activity. Methods: ANG II effects on [³H]-DA uptake and Na⁺,K⁺-ATPase were measured in samples from the outer renal cortex of Sprague-Dawley rats. Results: Inhibition of the phospholipase C (PLC) pathway blunted ANG II inhibitory effects on [³H]-DA uptake, since U-73122, 2-APB, TMB-8, chelerythrine and KN-93 (PLC, IP₃-dependent Ca²⁺ release channels, IP₃ receptors, protein kinase C and CaM kinase II inhibitors, respectively) each one blocked ANG II effects. Inhibition of adenylate cyclase pathway did not modify ANG II inhibitory effects on DA uptake. ANG II effects on [³H]-DA uptake were able to modify Na⁺,K⁺-ATPase activity in carbidopa-treated rats. Exogenous DA decreased while ANG II increased the enzyme activity. Neither the addition of DA together with ANG II, nor the extraneuronal DA uptake blocker hydrocortisone altered ANG II stimulatory effects on Na⁺,K⁺-ATPase activity, but hydrocortisone blocked the inhibitory effects of exogenous DA. Conclusion: Stimulation of renal AT₁ receptors by ANG II signals through the PLC pathway to inhibit extraneuronal DA uptake. DA and ANG II act through a common pathway involving reversible renal tubular Na⁺,K⁺-ATPase deactivation and activation, respectively. In addition, ANG II by itself is able to stimulate renal Na⁺,K⁺-ATPase activity.

Atrial natriuretic factor stimulates renal dopamine uptake mediated by natriuretic peptide-type A receptor.

To determine the effects of atrial natriuretic factor (ANF) on renal dopamine (DA) metabolism, 3H-DA and 3H-L-DOPA uptake by renal tubular cells was measured in experiments carried out in vitro in Sprague-Dawley rats. The receptor type involved was also analyzed. The results indicate that ANF increased at 30 min, DA uptake in a concentration-response fashion having 10 pM ANF as the threshold concentration. Conversely, the uptake of the precursor L-DOPA was not modified by the peptide. ANF effects were observed in tissues from external and juxtamedullar cortex and inner medulla. On this basis, 100 nM ANF was used to continue the studies in external cortex tissues. DA uptake was characterized as extraneuronal uptake, since 100 microM hydrocortisone blocked ANF-induced increase of DA uptake. Renal DA uptake was decreased at 0 degrees C and in sodium-free medium. The effects of ANF in these conditions were not present, confirming that renal DA uptake is mediated by temperature- and sodium-dependent transporters and that the peptide requires the presence of the ion to exhibit its actions on DA uptake. The biological natriuretic peptide type A receptor (NPR-A) mediates ANF effects, since 100 nM anantin, a specific blocker, reversed ANF-dependent increase of DA uptake. The natriuretic peptide type C receptor (NPR-C) is not involved, since the specific analogous 100 nM 4-23 ANF amide has no effect on renal DA uptake and does not alter the effects of 100 nM ANF. In conclusion, ANF stimulates DA uptake by kidney tubular cells. ANF effects are mediated by NPR-A receptors coupled to guanylate cyclase and cGMP as second messenger. The process involved was characterized as a typical extraneuronal uptake, and characterized as temperature- and sodium-dependent. This mechanism could be related to DA effects on sodium reabsorption and linked to ANF enhanced natriuresis in the kidney. The increment of endogenous DA into tubular cells, as a consequence of increased DA uptake, would permit D1 receptor recruitment and Na+,K+-ATPase activity inhibition, which results in decreased sodium reabsorption and increased natriuresis.

Angiotensin II Regulates Extraneuronal Dopamine Uptake in the Kidney

Background/Aims: Angiotensin II (ANG II) and dopamine (DA) are both important regulators of sodium and water transport across renal proximal tubules. Previous studies demonstrate that atrial natriuretic factor (ANF) can regulate renal DA uptake and thereby Na+,K+-ATPase activity in the external renal cortex. As ANG II counteracts most of the ANF biological effects, the aim of the present study was to evaluate ANG II effects on renal DA metabolism and identify the receptor involved. Methods: To determine ANG II effects on renal DA metabolism, we evaluated 3H-DA uptake in vitro in kidney tissue samples from Sprague-Dawley rats. Results: The results indicate that ANG II decreased DA uptake at 30 min, in a concentration-response fashion, in the external and juxtamedullar cortex. DA uptake was characterized as an extraneuronal uptake and decreased at 0°C and in sodium-free medium. The biological receptor type involved was AT1, since losartan reversed ANG II effects on DA uptake while AT2 receptors were not involved since PD 123319 did not affect ANG II effects. The absence of sodium did not alter the ANG II response. Conclusion: ANG II inhibits DA uptake by kidney tubular cells. These effects implicate AT1 receptors without participation of AT2 receptors. This mechanism could be related to the DA effects on sodium reabsorption and linked to ANG II antinatriuretic effects in the kidney.

Atrial natriuretic factor decreases renal dopamine turnover and catabolism without modifying its release

Atrial natriuretic factor (ANF) and dopamine (DA) are both important regulators of sodium and water transport across renal proximal tubules. Many evidences suggest that some of ANF inhibitory effects on sodium and water reabsorption are mediated by dopaminergic mechanisms. We have previously reported that ANF stimulates extraneuronal DA uptake in external renal cortex by activation of NPR-A receptors coupled to cGMP signal and PKG. Moreover, ANF enhanced DA-induced inhibition of Na(+)-K(+) ATPase activity. The aim of the present study was to evaluate if ANF could alter also renal DA release, catabolism and turn over. The results indicate that ANF did not affect basal secretion of the amine in external renal cortex or its KCl-induced release, but diminished DA turn over. Moreover, ANF diminished COMT and did not alter MAO activity. In conclusion, present results as well as previous findings show that ANF modifies DA metabolism in rat external renal cortex by enhancing DA uptake and decreasing COMT activity. All those effects, taken together, may favor DA accumulation into renal cells and increase its endogenous content and availability. This would permit D1 receptor recruitment and stimulation and in turn, Na(+), K(+)-ATPase activity over inhibition that results in decreased sodium reabsorption. Therefore, ANF and DA could act via a common pathway to enhance natriuresis and diuresis.

Effects of Angiotensin II on Renal Dopamine Metabolism: Synthesis, Release, Catabolism and Turnover

Background/Aims: Dopamine (DA) uptake inhibition in the renal cortex, elicited by angiotensin II (ANG II), is mediated by AT1 receptors and signals through the phospholipase C pathway and activation of protein kinase C and CaM-kinase II. By this indirect way, ANG II stimulates renal Na+,K+-ATPase activity through DA intracellular reduction. In the present work, we continued to study different aspects of renal DA metabolism in DA-ANG II interaction, such as DA synthesis, release, catabolism and turnover. Methods: ANG II effects on DA synthesis, release, catabolism and turnover were measured in samples from the outer renal cortex of Sprague-Dawley rats. Results: ANG II reduced renal aromatic acid decarboxylate activity without affecting basal secretion of DA or its KCl-induced release. Moreover, ANG II enhanced monoamine oxidase activity without altering catechol-o-methyl transferase activity and increased DA turnover. Conclusion: Current results as well as previous findings show that ANG II modifies DA metabolism in rat renal cortex by reducing DA uptake, decreasing DA synthesis enzyme activity and increasing monoamine oxidase activity, and DA turnover. Together, all these effects may reduce DA accumulation into renal cells and decrease its endogenous content and availability. This would prevent D1 receptor recruitment and stimulation, while diminishing DA inhibition of Na+,K+-ATPase activity and stimulating sodium reabsorption.

Effects of chronic fructose overload on renal dopaminergic system: alteration of urinary L-dopa/dopamine index correlates to hypertension and precedes kidney structural damage

Insulin resistance induced by a high-fructose diet has been associated to hypertension and renal damage. The aim of this work was to assess alterations in the urinary L-dopa/dopamine ratio over three time periods in rats with insulin resistance induced by fructose overload and its correlation with blood pressure levels and the presence of microalbuminuria and reduced nephrin expression as markers of renal structural damage. Male Sprague-Dawley rats were randomly divided into six groups: control (C) (C4, C8 and C12) with tap water to drink and fructose-overloaded (FO) rats (FO4, FO8 and FO12) with a fructose solution (10% w/v) to drink for 4, 8 and 12 weeks. A significant increase of the urinary L-dopa/dopamine ratio was found in FO rats since week 4, which positively correlated to the development of hypertension and preceded in time the onset of microalbuminuria and reduced nephrin expression observed on week 12 of treatment. The alteration of this ratio was associated to an impairment of the renal dopaminergic system, evidenced by a reduction in renal dopamine transporters and dopamine D1 receptor expression, leading to an overexpression and overactivation of the enzyme Na+, K+-ATPase with sodium retention. In conclusion, urinary L-dopa/dopamine ratio alteration in rats with fructose overload positively correlated to the development of hypertension and preceded in time the onset of renal structural damage. This is the first study to propose the use of the urinary L-dopa/dopamine index as marker of renal dysfunction that temporarily precedes kidney structural damage induced by fructose overload.

Atrial Natriuretic Peptide Stimulates Dopamine Tubular Transport by Organic Cation Transporters: A Novel Mechanism to Enhance Renal Sodium Excretion.

The aim of this study was to demonstrate the effects of atrial natriuretic peptide (ANP) on organic cation transporters (OCTs) expression and activity, and its consequences on dopamine urinary levels, Na+, K+-ATPase activity and renal function. Male Sprague Dawley rats were infused with isotonic saline solution during 120 minutes and randomized in nine different groups: control, pargyline plus tolcapone (P+T), ANP, dopamine (DA), D-22, DA+D-22, ANP+D-22, ANP+DA and ANP+DA+D-22. Renal functional parameters were determined and urinary dopamine concentration was quantified by HPLC. Expression of OCTs and D1-receptor in membrane preparations from renal cortex tissues were determined by western blot and Na+, K+-ATPase activity was determined using in vitro enzyme assay. 3H-DA renal uptake was determined in vitro. Compared to P+T group, ANP and dopamine infusion increased diuresis, urinary sodium and dopamine excretion significantly. These effects were more pronounced in ANP+DA group and reversed by OCTs blockade by D-22, demonstrating that OCTs are implied in ANP stimulated-DA uptake and transport in renal tissues. The activity of Na+, K+-ATPase exhibited a similar fashion when it was measured in the same experimental groups. Although OCTs and D1-receptor protein expression were not modified by ANP, OCTs-dependent-dopamine tubular uptake was increased by ANP through activation of NPR-A receptor and protein kinase G as signaling pathway. This effect was reflected by an increase in urinary dopamine excretion, natriuresis, diuresis and decreased Na+, K+-ATPase activity. OCTs represent a novel target that links the activity of ANP and dopamine together in a common mechanism to enhance their natriuretic and diuretic effects.