Shuji Arima

Aldosterone (Aldo) accelerates hypertension, proteinuria, and glomerulosclerosis in animal models of malignant hypertension or chronic renal failure. Aldo may exert these deleterious renal effects by elevating renal vascular resistance and glomerular capillary pressure. To test this possibility, directly examined were the action of Aldo on the afferent (Af) and efferent (Ef) arterioles (Arts). Examined were the effect of Aldo added to both the bath and lumen on the intraluminal diameter (measured at the most responsive point) of rabbits. Aldo caused dose-dependent constriction in both arterioles with a higher sensitivity in Ef-Arts. Vasoconstrictor action of Aldo was not affected by a mineralocorticoid receptor antagonist spironolactone and was reproduced by membrane-impermeable albumin-conjugated Aldo, suggesting that the vasoconstrictor actions are nongenomic. Pretreatment with neomycin (a specific inhibitor of phospholipase C) abolished the vasoconstrictor action of Aldo in both arterioles. In addition, the vasoconstrictor action of Aldo on Af-Arts was inhibited by both nifedipine and efonidipine, whereas that on Ef-Arts was inhibited by efonidipine but not nifedipine. The results demonstrate that Aldo causes nongenomic vasoconstriction by activating phospholipase C with a subsequent calcium mobilization thorough L- or T-type voltage-dependent calcium channels in Af- or Ef-Arts, respectively. These vasoconstrictor actions on the glomerular microcirculation may play an important role in the pathophysiology and progression of renal diseases by elevating renal vascular resistance and glomerular capillary pressure.

It has been reported that sensitivity to angiotensin II (Ang II) is higher in efferent (Ef) than afferent (Af) arterioles (Arts). We tested the hypothesis that this is due to arteriolar differences in the interaction between Ang II and endothelium-derived relaxing factor/nitric oxide (EDNO). Rabbit Af-Arts with glomerulus intact were microperfused in vitro at a constant pressure. Ef-Arts were perfused from the distal end of either the Af-Art (orthograde perfusion) or the Ef-Art (retrograde perfusion) to eliminate influences of the Af-Art or glomerulus, respectively. Ang II did not alter Af-Art luminal diameter until the concentration reached 10(-9) M, which decreased the diameter by 11 +/- 2.6% (n = 11; P < 0.002). In contrast, Ef-Arts became significantly constricted at concentrations as low as 10(-11) M with either perfusion. Surprisingly, the decrease in Ef-Art diameter at 10(-10), 10(-9), and 10(-8) M was significantly greater with retrograde perfusion (44 +/- 6.9%, 70 +/- 5.6%, and 74 +/- 4.1%, respectively; n = 5) than with orthograde perfusion (16 +/- 4.2%, 25 +/- 2.9%, and 35 +/- 3.5%; n = 9). ENDO synthesis inhibition with 10(-4) M nitro-L-arginine methyl ester (L-NAME) decreased the diameter to a greater extent in Af-Arts (22 +/- 3.0%; n = 11) compared to Ef-Arts with either orthograde (9.5 +/- 2.3%; n = 8) or retrograde perfusion (1.2 +/- 2.1%; n = 6). With L-NAME pretreatment, Af-Art constriction induced by 10(-10) M (14 +/- 4.0%, n = 9) and 10(-9) M Ang II (38 +/- 3.9%) was significantly greater compared to nontreated Af-Arts. In contrast, L-NAME pretreatment had no effect on Ang II-induced constriction in Ef-Arts with either perfusion. In conclusion, this study demonstrates higher sensitivity of Ef-Arts to Ang II, particularly with retrograde perfusion. Our results suggest that EDNO significantly modulates the vasoconstrictor action of Ang II in Af-Arts II but not Ef-Arts, contributing to the differential sensitivity to Ang II.

Angiotensin (A) II plays a critical role in vascular remodeling, and its action is mediated by type 1 AII receptor (AT1R). Recently, 15-deoxy-(Delta)(12,14)-prostaglandin J(2) and thiazolidinediones have been shown to be ligands for peroxisome proliferator-activated receptor (PPAR)-gamma and activate PPAR-gamma. In the present work, we have studied the effect of PPAR-gamma on AT1R expression in rat vascular smooth muscle cells (VSMCs). We observed that: 1) endogenous AT1R expression was significantly decreased by PPAR-gamma ligands both at messenger RNA and protein levels, whereas AT1R messenger RNA stability was not affected; 2) AII-induced increase of (3)H-thymidine incorporation into VSMCs was inhibited by PPAR-gamma ligands; 3) rat AT1R gene promoter activity was significantly suppressed by PPAR-gamma ligands, and PPAR-gamma overexpression further suppressed the promoter activity; 4) transcriptional analyses using AT1R gene promoter mutants revealed that a GC-box-related sequence within the -58/-34 region of the AT1R gene promoter was responsible for the suppression; 5) Sp1 overexpression stimulated AT1R gene transcription via the GC-box-related sequence, which was inhibited by additional PPAR-gamma overexpression; 6) electrophoretic mobility shift assay suggested that Sp1 could bind to the GC-box-related sequence whereas PPAR-gamma could not; 7) antibody supershift experiments using VSMC nuclear extracts revealed that protein-DNA complexes formed on the GC-box-related sequence, which were decreased by PPAR-gamma coincubation, were mostly composed of Sp1; and 8) glutathione S-transferase pull-down assay revealed a direct interaction between PPAR-gamma and Sp1. Taken together, it is suggested that activated PPAR-gamma suppresses AT1R gene at a transcriptional level by inhibiting Sp1 via a protein-protein interaction. PPAR-gamma ligands, thus, may inhibit AII-induced cell growth and hypertrophy in VSMCs by AT1R expression suppression and possibly be beneficial for treatment of diabetic patients with hypertension and atherosclerosis.

Although angiotensin II type 2 (AT2) receptor has recently been cloned, its functional role is not well understood. We tested the hypothesis that selective activation of AT2 receptor causes vasodilation in the preglomerular afferent arteriole (Af-Art), a vascular segment that accounts for most of the preglomerular resistance. We microperfused rabbit Af-Arts at 60 mmHg in vitro, and examined the effect of angiotensin II (Ang II; 10(-11)-10(-8) M) on the luminal diameter in the presence or absence of the Ang II type 1 receptor antagonist CV11974 (CV; 10(-8) M). Ang II was added to both the bath and lumen of preconstricted Af-Arts. Ang II further constricted Af-Arts without CV (by 74+/-7% over the preconstricted level at 10(-8) M; P < 0.01, n = 7). In contrast, in the presence of CV, Ang II caused dose-dependent dilation; Ang II at 10(-8) M increased the diameter by 29+/-2% (n = 7, P < 0.01). This dilation was completely abolished by pretreatment with an AT2 receptor antagonist PD123319 (10(-7) M, n = 6), suggesting that activation of AT2 receptor causes vasodilation in Af-Arts. The dilation was unaffected by inhibiting either nitric oxide synthase (n = 7) or cyclooxygenase (n = 7), however, it was abolished by either disrupting the endothelium (n = 10) or inhibiting the cytochrome P-450 pathway, particularly the synthesis of epoxyeicosatrienoic acids (EETs, n = 7). These results suggest that in the Af-Art activation of the AT2 receptor may cause endothelium-dependent vasodilation via a cytochrome P-450 pathway, possibly by EETs.