Abstract-The distribution of voltage-dependent calcium channels in kidney pre-and postglomerular resistance vessels was determined at the molecular and functional levels. Reverse transcription-polymerase chain reaction analysis of microdissected rat preglomerular vessels and cultured smooth muscle cells showed coexpression of mRNAs for T-type subunits (Ca V 3.1, Ca V 3.2) and for an L-type subunit (Ca V 1.2). The same expression pattern was observed in juxtamedullary efferent arterioles and outer medullary vasa recta. No calcium channel messages were detected in cortical efferent arterioles. Ca V 1.2 protein was demonstrated by immunochemical labeling of rat preglomerular vasculature and juxtamedullary efferent arterioles and vasa recta. Cortical efferent arterioles were not immunopositive.Recordings of intracellular calcium concentration with digital fluorescence imaging microscopy showed a significant increase of calcium in response to K ϩ (100 mmol/L) in isolated afferent arterioles (140Ϯ25%) and in juxtamedullary efferent arterioles (118Ϯ21%). These calcium responses were attenuated by the L-type antagonist calciseptine and by the T-type antagonist mibefradil. Intracellular calcium increased in response to K ϩ in cortical efferent arterioles (21Ϯ9%). Mibefradil and nickel concentration dependently blocked K ϩ -induced contraction of perfused rabbit afferent arterioles. Calciseptine blocked the contraction mediated by K ϩ (EC 50 8ϫ10 Ϫ14 ). S-(-)-Bay K 8644 had no effect on vascular diameter in the afferent arteriole. We conclude that voltage-dependent L-and T-type calcium channels are expressed and of functional significance in renal cortical preglomerular vessels, in juxtamedullary efferent arterioles, and in outer medullary vasa recta, but not in cortical efferent arterioles. Key Words: voltage-dependent calcium channel Ⅲ renal Ⅲ afferent and efferent arterioles T he largest drop in intravascular pressure in the kidney occurs in the glomerular arterioles and in the descending vasa recta in the kidney outer medulla. These microvessels are targets for humoral and nervous signals, which interact to determine arteriolar diameters and thereby regulate overall blood flow to the kidney and the distribution of blood flow between kidney regions. There is electrophysiological evidence for the presence of L-type voltage-gated calcium currents and also for T-type calcium currents in single vascular smooth muscle cells (VSMCs) of preglomerular vessels. 1,2 Also, excitation-contraction coupling in preglomerular renal vascular segments is highly dependent on calcium entry through voltage-dependent calcium channels. 3 For postglomerular vascular segments, conflicting data exist with respect to the involvement of voltage-gated pathways for calcium entry and contraction. 3-11 However, with some exceptions, 5,6,11 most studies have shown calcium influx pathways in efferent vessels that are not activated by depolarization 4,8 and are resistant to L-type calcium channel antagonists. 7,9,10 Single-cell calcium currents have not...

Aldosterone has been suggested to elicit vessel contraction via a nongenomic mechanism. We tested this proposal in microdissected, perfused rabbit renal afferent arterioles. Aldosterone had no effect on internal diameter in concentrations from 10(-10) to 10(-5) mol/L, but aldosterone abolished the ability of 100 mmol/L KCl to induce vascular contraction. The inhibitory effect of aldosterone was observed from 1 pmol/L. The inhibitory effect was significant after 5 minutes and maximal after 20 minutes and was fully reversible. Actinomycin D (10(-6) mol/L) prolonged the effect of aldosterone. The effect was abolished by the mineralocorticoid receptor antagonist spironolactone (10(-7) mol/L) but not by the glucocorticoid receptor antagonist mifepristone (10(-6) mol/L). The K+-mediated increase of intracellular calcium concentration in afferent arterioles was not affected by aldosterone. Mineralocorticoid receptor was detected by reverse transcription-polymerase chain reaction and immunohistochemistry in rat renal vasculature and rabbit endothelial cells. Inhibition of phosphatidylinositol (PI)-3 kinase with LY 294002 (3x10(-6) mol/L) restored sensitivity to K+ in the presence of aldosterone, and afferent arterioles were immunopositive for PI-3 kinase subunit p110alpha. Inhibition of NO formation by L-NAME (10(-4) mol/L) or inhibition of soluble guanylyl cyclase with 1H-(1,2,4)Oxadiazolo[4,3-a]quinoxaline-1-one restored K+-induced vasoreactivity in the presence of aldosterone. Similar to aldosterone, the NO donor sodium nitroprusside inhibited K+-induced vascular contraction. Geldanamycin (10(-6) mol/L), an inhibitor of heat shock protein 90, abolished aldosterone-induced vasorelaxation. We conclude that aldosterone inhibits depolarization-induced vasoconstriction in renal afferent arterioles by a rapid nongenomic mechanism that is initiated by mineralocorticoid receptor activation and involves PI-3 kinase, protein kinase B, and heat shock protein 90-mediated stimulation of NO generation.

Nitric oxide (NO) is an important negative modulator of tubuloglomerular feedback responsiveness. We recently found that macula densa expresses a-, b-, and g-splice variants of neuronal nitric oxide synthase 1 (NOS1), and NOS1b expression in the macula densa increases on a high-salt diet. This study tested whether upregulation of NOS1b expression in the macula densa affects sodium excretion and saltsensitive hypertension by decreasing tubuloglomerular feedback responsiveness. Expression levels of NOS1b mRNA and protein were 30-and five-fold higher, respectively, than those of NOS1a in the renal cortex of C57BL/6 mice. Furthermore, macula densa NO production was similar in the isolated perfused juxtaglomerular apparatus of wild-type (WT) and nitric oxide synthase 1a-knockout (NOS1aKO) mice. Compared with control mice, mice with macula densa-specific knockout of all nitric oxide synthase 1 isoforms (MD-NOS1KO) had a significantly enhanced tubuloglomerular feedback response and after acute volume expansion, significantly reduced GFR, urine flow, and sodium excretion. Mean arterial pressure increased significantly in MD-NOS1KO mice (P,0.01) but not NOS1flox/flox mice fed a high-salt diet. After infusion of angiotensin II, mean arterial pressure increased by 61.6 mmHg in MD-NOS1KO mice versus 32.0 mmHg in WT mice (P,0.01) fed a high-salt diet. These results indicate that NOS1b is a primary NOS1 isoform expressed in the macula densa and regulates the tubuloglomerular feedback response, the natriuretic response to acute volume expansion, and the development of salt-sensitive hypertension. These findings show a novel mechanism for salt sensitivity of BP and the significance of tubuloglomerular feedback response in long-term control of sodium excretion and BP.

Abstract. Adenosine induces vasoconstriction of renal afferent arterioles through activation of A1 adenosine receptors (A1AR). A1AR are directly coupled to Gi/Go, resulting in inhibition of adenylate cyclase, but the contribution of this signaling pathway to smooth muscle cell activation is unclear. In perfused afferent arterioles from mouse kidney, adenosine and the A1 agonist N 6