1. ¹Department of Internal Medicine II, Yokohama City University, Yokohama,
2. ²Lead Generation Research Laboratories, Tanabe Seiyaku, Kashima, Osaka,
3. ³Department of Neuroanatomy, Yokohama City University, Yokohama,
4. ⁴the Institute of Applied Biochemistry, University of Tsukuba, Tsukuba, Japan

Correspondence: Minoru Kihara, Department of Internal Medicine II, Yokohama City University School of Medicine, Kanazawa-ku, Yokohama 236, Japan.

Received 5 September 1997; Revised 30 December 1997; Accepted 2 January 1998.

Abstract

Expression of neuronal type nitric oxide synthase and renin in the juxtaglomerular apparatus of angiotensin type-1a receptor gene-knockout mice. Angiotensin type-1a (AT1a) receptor gene-knockout (AT1a^-/-) mice exhibit chronic hypotension and renin overproduction. In the kidneys of AT1a^-/- mice, the activity of neuronal type nitric oxide synthase (N-NOS) was histochemically detected by nicotinamide adenine dinucleotide phosphate (NADPH) diaphorase (NADPHd) reaction combined with N-NOS immunohistochemistry. The localization of renin was detected by immunohistochemistry and the results were analyzed morphometrically. The levels of N-NOS and renin mRNA in the renal cortical tissue were determined by reverse transcription-PCR and Northern blot analysis, respectively. In the renal sections from wild-type mice, NADPHd activity and N-NOS immunoreactivity were localized to the discrete region of the macula densa in contact with the parent glomerulus. In contrast, N-NOS-positive macula densa cells were distributed beyond the original location of the macula densa, occasionally extending to the opposite side of the distal tubules. The mean number of N-NOS positive macula densa cells was significantly increased in AT1a^-/- mice (186 per 100 glomeruli) compared with wild-type mice (65 per 100 glomeruli). AT1a^-/- mice showed 1.4-times higher N-NOS mRNA levels in the renal cortical tissues than wild-type mice. The plasma renin activity was significantly higher in AT1a^-/- mice (205.5 26.1 ng/ml/hr) than in wild-type mice (8.0 0.2 ng/ml/hr). The renin-positive areas per glomerulus and renal renin gene expression were 12-times and 2.6-times higher in AT1a^-/- mice than in wild-type mice, respectively. These abnormalities, however, were less remarkable in AT1a^-/- mice compared with angiotensinogen-knockout mice. When AT1a^-/- mice were fed a high-salt diet, the signal intensity of the NADPHd reaction and the number of positively-stained macula densa cells were significantly decreased. The levels of renal cortical N-NOS mRNA were also suppressed by the treatment. Dietary salt loading produced a parallel decrease in plasma renin activity, renal renin-immunoreactive areas, and the levels of renin mRNA without affecting systemic blood pressure. These results provide evidence for the possible involvement of N-NOS at the macula densa in the increased renin production in AT1a^-/- mice.