Renin-containing Müller cells of the retina display endocrine features.

PURPOSE: An ocular renin-angiotensin system has been implicated in the proliferation of retinal blood vessels and blindness in diabetes mellitus. Its cellular basis has not been established. The objective was to identify sites of renin synthesis, secretion, and processing in eyes from humans, BALB/c mice, Sprague-Dawley rats, and a hypertensive transgenic rat model (mREN-2) that displays amplified extrarenal renin synthesis. METHODS: Paraffin sections of eyes were incubated with antisera to renin protein, prorenin, vimentin, and Müller cells. Enzyme kinetic renin assay was performed on extracts of whole eyes (excluding lens and vitreous) and comparisons made with adrenal glands and kidneys. For detection of renin mRNA, retinas were separately pooled from BALB/c and Swiss mice. RESULTS: In normal rodent and autopsy human eyes, labeling for renin, vimentin, and Müller cell protein was observed in the cytoplasm of all macroglial Müller cells, with renin labeling most obvious in endfeet closely apposed to retinal blood vessels. Prorenin labeling was not detected. Less intense renin labeling, again without prorenin, was seen in nonpigmented ciliary epithelium of rodents. In transgenic (mREN-2) rat eyes, renin and prorenin labeling of Müller cells and nonpigmented ciliary epithelium were intense. Prorenin was localized to the posterior region of Müller cells but only sparsely to endfeet in rodent retinas, and renin was present only in an active form in amounts one third that of one adrenal. Renin mRNA was readily detected. In human retina, renin was present in active and pro-forms, and the total amount was approximately one fiftieth that of adrenal. CONCLUSION: Renin is synthesized in the retina and is specifically localized to the macroglial Müller cells. Nonpigmented ciliary epithelium also contains renin. The presence of prorenin in the posterior part of the Müller cell, with active renin throughout but notably in endfeet in apposition to retinal capillaries, suggests directional processing of renin. These findings are consistent with earlier suggestions that retinal neovascularization may be associated with Müller cell dysfunction.

The use of non-invasive blood pressure measurements to measure pressor responses in rats during air stress.

1. A non-invasive tail cuff method was validated against direct intra-arterial blood pressure measurements (r2 greater than 0.9) and then used to measure the systolic blood pressure (SBP) responses to air-jet stimulation in mature conscious Sprague-Dawley (SD) and spontaneously hypertensive rats (SHR). The experiments with the SHR were conducted in parallel with age matched SD rats. 2. All rats were trained to remain in perspex holders and when their SBP had stabilized, a jet of air of 10 min duration was directed at the rat's nose via rubber tubing. The blood pressure was measured during the first, fifth and tenth minute. 3. The maximum SBP response normally occurred during the first min of air-jet stimulation and averaged +30 mmHg in the SHR and +21 mmHg in the SD. Although of higher magnitude in the SHR, when expressed as a percentage of the control SBP, the rise was similar for both strains, 13% and 14%, respectively. 4. The pressor response of three SD rats stimulated daily for up to 12 days did not show any evidence of habituation to the stimulus. 5. The pressor responses to air-jet stimulation were abolished in anaesthetized rats, suggesting that they are primarily higher centre responses.

Comparison of the actions of the angiotensin-converting enzyme inhibitors enalapril and S-9490-3 in sodium-deplete and sodium-replete spontaneously hypertensive rats.

The hypotensive action of the angiotensin-converting enzyme (ACE) inhibitors enalapril and S-9490-3 was examined in conscious, chronically cannulated Na+-replete and Na+-deplete spontaneously hypertensive rats (SHR) of the Okamoto strain. Blood pressure, plasma ACE activity, plasma renin activity (PRA), and pressor responses to intravenous bolus injections of angiotensin I (AI) were measured over a 24-h period following a single oral dose of ACE inhibitor (0.3, 1.0, and 3.0 mg/kg) or vehicle. S-9490-3 caused a significantly greater hypotensive response and inhibition of plasma ACE and AI pressor responses than enalapril for each dose in both diet groups. Single oral doses of both drugs (3 mg/kg) caused slow, progressive falls in blood pressure which were maximal at 12 h. In contrast, inhibition of plasma ACE was maximal 1 h following the oral dose and returned to control levels over the 24-h period. The inhibition of the pressor response to intravenous AI paralleled, and was significantly correlated with, the inhibition of plasma ACE. There was no correlation between the maximal fall in blood pressure with PRA or with inhibition of plasma ACE activity in either diet group. The hypotensive response to both drugs at the 3-mg/kg dose was greater in Na+-deplete SHR than in Na+-replete animals. Both drugs caused large rises in PRA. The ACE inhibitor S-9490-3 is a significantly more potent hypotensive agent than enalapril in the SHR and a significantly more potent ACE inhibitor in vivo. The hypotensive response to both drugs was dissociated in onset and duration from the inhibition of plasma ACE and AI pressor responses.

Sodium detection threshold and preference for sodium chloride in humans on high and low sodium diets.

Sodium detection threshold and preference for sodium chloride was examined in human volunteers on 2 week periods of high and low salt diets. There was no significant difference in salivary electrolytes or sodium detection threshold between the high and low salt diet periods. Sodium preference, defined as the final sodium concentration of unsalted tomato juice following the ad libitum addition of NaCl, was significantly less on the low salt diet when compared to the high salt period. These results suggest that short periods of dietary salt modification can result in marked shifts in preference for NaCl in the absence of changes in the physiological parameters measured.

Vascular prostacyclin and Goldblatt hypertensive rats.

Vascular prostacyclin production in Goldblatt hypertension was examined in one-kidney, one clip (1K, 1C) and two-kidney, one clip (2K, 1C) rat models. Vasodepressor responses to prostacyclin and nitroprusside correlated well with resting blood pressure in both groups of rats, but when measured as a percentage of resting blood pressure the responses did not differ significantly between hypertensive rats and the normotensive controls within each group. In contrast, the vasodepressor effects of arachidonic acid (1-3 mg/kg, i.v.) were much greater in the 1K, 1C rats than in their normotensive controls, but did not differ significantly between hypertensive 2K, 1C rats and sham-operated controls. The effects of arachidonic acid were virtually abolished by indomethacin (10 mg/kg, i.v.). The metabolism of [14C]-arachidonic acid was also studied in isolated aortae of both one- and two-kidney rats by high pressure liquid chromatography of extracts of the incubation mixture. [14C]-6-oxo-PGF1 alpha was the only prostanoid conversion product recovered from the incubations and significantly more of this metabolite was produced by aortic tissue from 1K, 1C rats than from normotensive controls. There was no difference in [14C]-6-oxo-PGF1 alpha production between 2K, 1C rats and controls. These results demonstrate an enhanced ability of vascular tissue from 1K, 1C hypertensive rats to convert exogenous arachidonate to vasodilator prostacyclin, but this is not evident in the two-kidney model. Although enhanced biosynthetic capacity for prostacyclin in the one-kidney model and spontaneously hypertensive rats does not lessen peripheral vascular resistance, it might reflect a fundamental disturbance in phospholipid metabolism which contributes to increased vascular resistance.

Sodium intake and exchangeable sodium in hypertensive rats.

Exchangeable sodium, extracellular fluid volume and sodium intake were significantly higher in spontaneously hypertensive rats (SHR) than in normotensive controls. The increased exchangeable sodium was due to the expanded extracellular fluid sodium content. Two kidney one clip Goldblatt renal hypertensive rats develop thirst and high renin levels. When drinking saline the rats have markedly increased exchangeable sodium levels, but those drinking water did not. It is proposed that the exaggerated saline intake leads to a volume dependent component of the hypertension in both models. In the SHR both the salt appetite and the hypertension may be induced by angiotensin acting via central mechanisms.

Evidence for a functional central dopaminergic insufficiency in the spontaneously hypertensive rat.

Several neuroendocrine abnormalities reported in the spontaneously hypertensive rat (SHR) suggest that the function of the dopaminergic tuberoinfundibular tract is impaired in this strain. SHR were pretreated with the peripheral acting inhibitor of dopa decarboxylase, carbidopa, or vehicle. Intravenous injection of L-dopa, the precursor of dopamine, lowered blood pressure only in the carbidopa treated SHR suggesting a central dopamine action. Intracerebroventricular injection of dopamine in SHR lowered blood pressure towards normal. The data are in agreement with the hypothesis that SHR have a functional central dopaminergic insufficiency.

Failure of captopril to lower blood pressure in spontaneously hypertensive rats offered water and saline to drink.

The hypotensive action of chronic oral captopril treatment (50 mg/kg per day) was examined in two groups of male spontaneously hypertensive rats (SHR) of the Okamoto strain, one group offered a choice of water and 0.9% saline as drinking fluid, the other offered water alone, a third group of SHR, offered a choice of water and 0.9% saline, were treated with vehicle (0.9% saline, 2 ml/kg per day). Captopril treatment, over ten days, significantly lowered blood pressure in the group drinking water only but failed to significantly alter the blood pressure of SHR drinking a choice of water and 0.9% saline. Vehicle treatment did not alter the blood pressure of SHR drinking a choice of saline and water. In an identical experiment using male, genetically hypertensive rats (GHR) of the Smirk strain, captopril lowered blood pressure to the same extent in GHR drinking either a choice of water and 0.9% saline or water alone. In conclusion, the reported exaggerated saline preference of the spontaneously hypertensive rat appears to antagonize the hypotensive action of captopril.

Modulation of brain angiotensin-converting enzyme by dietary sodium and chronic intravenous and intracerebroventricular fusion of angiotensin II.

Angiotensin-converting enzyme (ACE) in rat brain closely resembled that in lung in its kinetics with the substrate Hip-His Leu, the inhibitors SQ 20,881 and SQ 14,225, and iun its Cl- activation profile. Modification of dietary NaCl intake was associated with marked changes in brain ACE activity. Sodium-loaded rats had lower activity of ACE in hypothalamus, striatum, and midbrain, and higher activity in spinal cord compared to controls. In sodium-restricted rats, ACE was elevated in pituitary and depressed in spinal cord. Chronic intravenous infusion of angiotensin (AII) was associated with a pattern of changes partly resembling sodium loading: ACE was depressed in hypothalamus and striatum but elevated in midbrain. After chronic intracerebroventricular infusion of AII, ACE was elevated in striatum and hippocampus, and depressed in spinal cord; a pattern of changes quite different from those associated with intravenous AII. These results show that ACE in several brain regions is sensitive to dietary sodium intake and support the hypothesis that angiotensin-containing neurons in these areas might be responsive to NaCl status of the animal. The observed changes in brain ACE do not seem to be explained in any simple manner by changes in circulating or central angiotensin II.

Impaired angiotensin-induced drinking in spontaneously hypertensive rats.

The drinking response during the 30 minutes following intracerebro-ventricular injection of angiotensin II (AII) (1 to 200 ng) was compared in male spontaneously hypertensive rats (SHR) and normotensive Wistar Kyoto control rats (WK). SHR drank significantly less than WK at the 10, 50 and 100 ng doses. In contrast, responses to intracerebroventricular carbachol and intraperitoneal hypertonic saline were not different between SHR and WK. These agents are believed to induce drinking by mechanisms independent of angiotensin. Binding of I125-labelled AII to particles prepared from the hypothalamus, thalamus, septum and midbrain (HTSM) region was measured in one month old male and two month old female SHR and their respective age matched WK controls. No differences were found in binding between SHR and WK of either sex. These data demonstrate an impairment of drinking responses in the SHR which seem to be specific for angiotensin. This finding supports the hypothesis that the CNS angiotensin system might be abnormal in these animals.

Exaggerated salt appetite of spontaneously hypertensive rats is decreased by central angiotensin-converting enzyme blockade.

Injection of angiotensin II (AII) into the cerebral ventricles at doses as low as 1 pmol h-1 results in a marked stimulation of salt and water ingestion in the rat. Evidence that AII is produced in the central nervous system independently of the circulating renin-angiotensin system (RAS) raises the possibility that endogenous brain AII is involved in the physiological regulation of thirst. The role of brain AII in salt appetite is still unclear. Here we confirm that the spontaneously hypertensive rat (SHR), believed to have elevated levels of brain AII, possesses an exaggerated salt appetite compared with its normotensive controls. We also show that this exaggerated salt appetite is reduced by chronic central treatment with the angiotensin-converting enzyme inhibitor, captopril, while that of the normotensive controls is unaffected. Our study suggests that a central neuropeptide, probably AII, is involved in the maintenance of the exaggerated salt appetite in this model of hypertension.

In vivo comparison of three orally active inhibitors of angiotensin-converting enzyme.

This study was designed to compare the activity of three structurally different drugs (SQ 14225 [captopril], SA 446 and MK 421) as inhibitors of angiotensin I-converting enzyme in vivo and to compare their effects on blood pressure in spontaneously hypertensive rats and one clip, two kidney renal hypertensive rats. All the three drugs were potent, orally effective converting enzyme inhibitors. The relative durations of their actions as inhibitors of angiotensin-converting enzyme, from longest to shortest, were as follows: MK 421, SQ 14225 and SA 446. MK 421 appears the most potent on a molar basis. In renal hypertensive rats the drugs appeared equipotent, although the duration of action of MK 421 was prolonged and SA 446 shorter than that of SQ 14225. SA 446 was less effective than the other two compounds in spontaneously hypertensive rats.

Immunoreactive beta-endorphin levels in plasma and pituitary tissue from genetically hypertensive and normotensive rats.

1. Levels of immunoreactive beta-endorphin were measured in neurointermediate lobes, anterior lobes and plasma from the Japanese and New Zealand strains of genetically hypertensive rats and their normotensive controls. 2. No significant differences were observed in beta-endorphin between the hypertensive and normotensive rats of the New Zealand strain. 3. The hypertensive rats of the Japanese strain showed significantly higher levels of beta-endorphin in neurointermediate lobe and lower levels in plasma than their normotensive controls. 4. These results suggest that the differences in beta-endorphin levels in the Japanese strain reflect a genetic difference not necessarily related to elevated blood pressure.

Failure of aspirin to modify the hypotensive action of captopril in spontaneously hypertensive rats.

1. Oral administration of the angiotensin converting enzyme inhibitor, captopril (30 mg/kg per day) to spontaneously hypertensive rats of the Okamoto strain progressively reduced arterial blood pressure by 60 mmHg over 4-5 days. 2. Oral treatment of spontaneously hypertensive rats with aspirin (200 mg/kg per day) for one week did not alter blood pressure, but it greatly reduced the vasodepressor effects of intravenous injections of arachidonic acid (3 mg/kg). 3. The fall in blood pressure of spontaneously hypertensive rats treated concurrently with both aspirin (200 mg/kg per day) and captopril (30 mg/kg per day) was not different to the fall observed in rats treated with captopril alone. 4. The hypotensive action of captopril in spontaneously hypertensive rats does not appear to be due to stimulation of vasodilator prostanoid biosynthesis.