ENHANCEMENT OF ENDOTHELIUM-DEPENDENT RELAXATION IN THE AORTA FROM STROKE-PRONE SPONTANEOUSLY HYPERTENSIVE RATS AT DEVELOPMENTAL STAGES OF HYPERTENSION

1. Endothelium-dependent relaxation (EDR) in aortic rings from young (8 weeks) and adult (16 weeks and 20 weeks) stroke-prone spontaneously hypertensive rats (SHRSP) was investigated in comparison with age-matched Wistar-Kyoto rats (WKY). 2. At 8 weeks, acetylcholine (3×10^?-⁹-^?10^?-⁵ mol/L) and ionomycin (4×10^?8-10^?6 mol/L)-induced EDR in SHRSP aortae was significantly enhanced compared to that in WKY aortae. Mechanical denudation of the endothelium completely abolished, and pretreatment of aortae with N^G-monomethyl L-arginine (1 mmol/L), an inhibitor of nitric oxide formation, greatly reduced the relaxation in both strains. Indomethacin (10^?5 mol/L), a cyclo-oxygenase inhibitor that blocks the production of endothelium-derived contracting factors, did not significantly alter the relaxation by acetylcholine at this age. There was no difference in endothelium-independent relaxation of denuded aortae by sodium nitroprusside (10^?9-10^?6 mol/L) and 8-bromoguanosine 3‘, 5‘-cyclic monophos-phate (10^?6-10^?3 mol/L). 3. In adult SHRSP with established hypertension, however, the acetylcholine (10^?8-^?10^?5 mol/L)-induced relaxation markedly diminished at any of the concentrations tested compared to that observed in 8 week old SHRSP and WKY at 8–20 weeks of age. This finding differed from other observations where the relaxation in SHRSP was impaired only at higher concentrations of acetylcholine. Indomethacin pretreatment of aortae from 20 week old SHRSP restored acetylcholine-induced EDR to a level comparable with that in age-matched WKY. 4. These results suggest that the aorta from young SHRSP releases more endothelium-derived relaxing factors in response to acetylcholine and ionomycin, but the relaxation greatly diminishes in adult SHRSP with established hypertension. This may be due to counteraction of endothelium-derived contracting factors released from the endothelium with functional changes resulting from the long duration of the hypertension.     

EFFECT OF TRANSFORMING GROWTH FACTOR-β1 ON PLATELET-DERIVED GROWTH FACTOR RECEPTOR BINDING AND GENE EXPRESSION IN VASCULAR SMOOTH MUSCLE CELLS FROM SHR AND WKY RATS

1. This study examined the effects of transforming growth factor-βl (TGF-β1) on platelet-derived growth factor-BB (PDGF-BB)-stimulated DNA synthesis, [¹²⁵I]-PDGF-BB receptor binding and PDGF-β receptor mRNA expression in vascular smooth muscle cells (VSMC) isolated from Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR). 2. TGF-β1 inhibited by 40% DNA synthesis stimulated by PDGF-BB in VSMC from WKY rats but potentiated by 20% growth factor-stimulated DNA synthesis in VSMC from the SHR. 3. Since the difference in effect of TGF-β1 could not be attributed to differential regulation of [¹²⁵I]-PDGF-BB binding activity and PDGF-β receptor mRNA expression, it is suggested that alterations in intracellular signalling pathways may account for the differential effects of TGF-β1 on PDGF-BB-stimulated DNA synthesis in VSMC from SHR and WKY rats.     

MULTIPLE GROWTH ABNORMALITIES IN VASCULAR SMOOTH MUSCLE FROM SPONTANEOUSLY HYPERTENSIVE RATS

1. In tissue culture the growth characteristics of aortic smooth muscle cells isolated from spontaneously hypertensive rats (SHR) were compared with those of normotensive Wistar-Kyoto (WKY) rats. 2. Aortic smooth muscle cells from SHR exhibit enhanced proliferation when grown in the presence of low (1%) and moderate (5%) concentrations of fetal calf serum. 3. Cell quiescence in cultures of smooth muscle from SHR becomes apparent at cell densities approximately 20% higher than in cultures from WKY rats. 4. These different growth characteristics of smooth muscle between the two strains of rats may contribute to the early pre-hypertensive development of vascular hypertrophy in the SHR.     

IMMUNOREACTIVE β-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.     

DIFFERENTIAL REGULATION BY TRANSFORMING GROWTH FACTOR-β1 OF PLATELET-DERIVED GROWTH FACTOR- STIMULATED PROLIFERATION OF VASCULAR SMOOTH MUSCLE CELLS FROM SHR AND WKY RATS

1. This study has examined and compared the time-course of action of transforming growth factor-beta 1 (TGF-beta 1) on platelet-derived growth factor-BB-stimulated proliferation of vascular smooth muscle cells isolated from normotensive Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR). 2. Transforming growth factor-beta 1 inhibited vascular smooth muscle cell proliferation stimulated by platelet-derived growth factor-BB in WKY rats by approximately 60-75%. 3. In contrast, transforming growth factor-beta 1 potentiated an 8-35% growth factor action on cell proliferation in the SHR. 4. Defects in transforming growth factor-beta 1 action may be part of the molecular mechanism responsible for the development of vascular hypertrophy in the SHR.     

VASCULAR SMOOTH MUSCLE CELL PROLIFERATION IN SHR AND WKY RATS: EVIDENCE FOR SPECIFIC DIFFERENCES IN GROWTH INHIBITORY REGULATORY MECHANISMS

1. This study examined and compared the actions of transforming growth factor-β₁ (TGF-β₁), heparin, dexamethasone and interferon-γ on platelet-derived growth factor-BB (PDGF-BB)-stimulated proliferation of vascular smooth muscle cells (VSMC) from normotensive, Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR). 2. Heparin, dexamethasone and interferon-γ all inhibited VSMC proliferation stimulated by PDGF-BB in both SHR and WKY rats. There was no difference (P>0.05) in their inhibitory effects, which varied between 40 and 85% for the different agents. 3. Similarly, TGF-β₁ inhibited PDGF-BB-stimulated VSMC proliferation in WKY rats by approximately 50%. In contrast, TGF-β₁ potentiated growth factor action on cell proliferation in the SHR by approximately 40%. 4. Specific TGF-β₁-stimulated regulatory mechanisms involved in the inhibition of proliferation are absent in SHR and this defect may contribute to the vascular hypertrophy which is apparent in genetic hypertension.     

ALTERED LEVELS OF NEUROPEPTIDES IN THE MEDULLA AND SPINAL CORD OF SPONTANEOUSLY HYPERTENSIVE RATS

1. Spontaneously hypertensive rats (SHR) are useful for investigating the possible pathophysiological and neurochemical basis of human essential hypertension. 2. The accepted pathogenic mechanism of hypertension in SHR is an increased central sympathetic drive which results in an increased peripheral resistance. 3. The neurochemical basis of the increased sympathetic drive is unknown. The observation that there are reduced levels of neuropeptides (vasoactive intestinal peptide, neuropeptide Y, cholecystokinin octapeptide, neurotensin and calcitonin gene related peptide) in the spinal cord in SHR rats compared with age and gender matched Wistar-Kyoto normotensive rats could provide a basis for understanding the mechanism of hypertension in SHR.     

DIFFERENCES IN ENDOTHELIUM-DEPENDENT RELAXATION IN VARIOUS ARTERIES FROM WATANABE HERITABLE HYPERLIPIDAEMIC RABBITS WITH INCREASING AGE

1. Endothelium-dependent relaxation in response to acetylcholine (ACh) and the calcium ionophore A 23187 was examined in aorta, coronary, basilar and renal arteries isolated from Watanabe heritable hyperlipidaemic (WHHL) rabbits of 2, 6 and 12 months of age, with normolipidaemic heterozygous WHHL rabbits as controls. 2. In the rings of WHHL rabbit aortae and coronary arteries preconstricted with vasoconstrictors, endothelium-dependent relaxation in response to ACh was attenuated with age compared to the heterozygous WHHL rabbits. A significant negative correlation was found between the total cholesterol content and the relaxation response to ACh in the aortae or coronary arteries from 6 and 12 month old WHHL rabbits. 3. In the rings of basilar arteries, endothelium-dependent relaxations to ACh were not modified with age. Similarly, in the rings of renal arteries, the relaxation response to ACh was not changed with age, but in the 6 and 12 month preparations, after the age of 6 months, a contraction following the relaxation appeared at higher concentrations of ACh (10^?7 to 10^?6 mol/L). The contraction was endothelium-dependent and inhibited by indomethacin. 4. A 23187-induced endothelium-dependent relaxations were also markedly attenuated in the aorta and significantly in the coronary artery with age. 5. Endothelium-independent relaxation to sodium nitroprusside was not changed in all arteries from WHHL rabbits of different ages. 6. These findings indicate that in the aorta and coronary artery of the WHHL rabbit, the endothelium-dependent relaxation to ACh and A 23187 becomes impaired with increasing age (i.e., with the progression of cholesterol deposition in the arterial wall) but is preserved in the basilar and renal artery.     

RELATIONSHIP BETWEEN NIFEDIPINE SENSITIVITY OF AORTAE AND BLOOD PRESSURE OF STROKE-PRONE SPONTANEOUSLY HYPERTENSIVE RATS

1. We have previously described an increased sensitivity to inhibition by nifedipine of noradrenaline-induced contractures of blood vessels in hypertension. In this study we have investigated whether changes in blood pressure (BP) change the sensitivity to nifedipine and K⁺ of aortic rings from normotensive (Wistar-Kyoto rats, WKY) and stroke-prone spontaneously hypertensive rats (SHRSP). 2. SHRSP were treated with: hydralazine plus hydrochloro-thiazide; captopril plus hydrochlorothiazide; hydralazine plus guanethidine; or captopril alone. WKY rats were treated with deoxycorticosterone acetate (DOCA) and NaCl. Treatment commenced from 5 weeks of age and continued until 13–15 weeks. 3. The SHRSP treatments produced similar reductions in BP, and the BP of all the treated groups were significantly lower than the mean BP of untreated SHRSP (201.0 ± 7.7 mmHg). The mean BP of the treated WKY rats (134.2 ± 7.6 mmHg) was significantly higher than the mean BP of the untreated WKY rats (86.8 ± 7.4 mmHg). 4. An area-under-curve (AUC) analysis of the inhibitory effects of nifedipine on responses of aortae to noradrenaline showed no differences between treated and untreated SHRSP groups (overall mean 40.6 ± 1.9% and 43.4 ± 3.4% inhibition of control AUC, respectively), or between DOCA-salt treated WKY and untreated WKY groups (58.8 ± 5.9 and 64.8 ± 2.3, respectively). Noradrenaline-induced contractures of aortae from all SHRSP groups were significantly more sensitive to inhibition by nifedipine than aortae from both WKY groups. 5. The molar concentration of agonist required to evoke 50% of the maximum response (EC₅₀) values for potassium chloride (KCI) were significantly increased in the aortae of all treated SHRSP groups in comparison to those from untreated SHRSP (treated SHRSP groups, 15.53 ± 0.68 mmol/L vs untreated SHRSP group, 11.36 ± 1.10 mmol/L). The EC₅₀ values for KC1 for the aortae from the DOCA-treated WKY rats were significantly less than those from aortae of the untreated WKY (11.80 ± 0.80 and 17.08 ± 1.50 mmol/L, respectively). 6. We conclude that reduction (in SHRSP) or increase (in WKY) of the BP has no effect on the sensitivity of aortic smooth muscle to the inhibitory effects of nifedipine on responses to noradrenaline, suggesting that alterations in voltage-dependent Ca²⁺ mechanisms may be a primary phenomenon in the SHRSP. In contrast, the fact that sensitivity to KC1 changes in the treated SHRSP and WKY aortae suggests such sensitivity is secondary to the BP and thus a separate phenomenon from voltage-dependent Ca²⁺ mechanisms.     

THE RELATIONSHIP BETWEEN TISSUE LEVELS OF CYCLIC GMP AND TRACHEAL SMOOTH MUSCLE RELAXATION IN THE GUINEA-PIG

The effect of cyclic GMP was investigated using guinea-pig tracheal smooth muscle. 8-Bromo-cyclic GMP showed a dose-dependent relaxation of spontaneous tension of tracheal smooth muscle. Administration of sodium nitroprusside induced dose-dependent relaxation of tracheal smooth muscle as well as an increase in tissue levels of cyclic GMP. Nicorandil, N-(2-hydroxyethyl) nicotinamide nitrate showed dose-dependent relaxation of tracheal smooth muscle and an increase in cyclic GMP levels in the tissue. N-(2-aminoethyl) nicotinamide dihydrochloride, which is a nicorandil derivative and differs minimally in its molecular structure (-NH2 vs -NO2), had neither a relaxant effect on tracheal smooth muscle nor did it increase the level of cyclic GMP in the tissue. The rise cyclic GMP levels preceded the relaxation of tracheal smooth muscle induced by sodium nitroprusside. These results suggest that cyclic GMP is one of the relaxant factors and that nitro-derivatives exhibit their relaxant effect on the smooth muscle mediated by an increase in cyclic GMP level.     

AUGMENTATION OF ANGIOTENSIN II RELEASE FROM ISOLATED MESENTERIC ARTERIES OF WISTAR-KYOTO AND SPONTANEOUSLY HYPERTENSIVE RATS FOLLOWING NEPHRECTOMY

1. We previously reported that angiotensin II release from the mesenteric arteries of Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR) increased in a time-dependent manner as a result of the isolation of the arteries and perfusion. This phenomenon appeared to be due to the withdrawal of circulating angiotensin II (AII). 2. The purpose of the present study was to test the hypothesis that vascular AII generation may be negatively regulated by circulating AII in WKY and SHR, and to clarify the role of this vascular angiotensin II in the sustained hypertension of SHR following nephrectomy. 3. The mesenteric arteries from kidney-intact and nephrectomized WKY and SHR were perfused and the amount of AII released into the perfusate was measured. The effects of the angiotensin converting enzyme inhibitor, captopril, and the effects of supplementation of renal renin and circulating angiotensins to nephrectomized rats, by blood exchange between kidney-intact and nephrectomized rats, on AII release were examined to clarify the pathway of vascular AII generation after nephrectomy. 4. Nephrectomy caused augmentation of vascular AII release both in WKY and SHR in spite of the abolishment of circulating renin. Captopril reduced this enhanced release of AII, but blood exchange did not affect it. There was no significant difference in these responses between WKY and SHR. 5. These results suggest that WKY and SHR have in common a potent pathway for production of vascular AII in response to the withdrawal of circulating AII, although this pathway is not responsible for the sustained hypertension of SHR after nephrectomy. The precise pathophysiological role of this pathway remains to be elucidated.     

EFFECT OF ANGIOTENSIN-CONVERTING ENZYME INHIBITORS CAPTOPRIL AND ENALAPRIL ON cAMP CONTENT OF SPECIFIC BRAIN AREAS IN SPONTANEOUSLY HYPERTENSIVE RATS

1. The influence of two angiotensin-converting enzyme inhibitors, captopril and enalapril, on the cAMP content of microdissected brain areas was examined in spontaneously hypertensive rats. Both drugs depleted systolic arterial blood pressure significantly. 2. Captopril and enalapril increased the level of cAMP in catecholaminergic cell groups in the lower brain-stem. Captopril was more effective in the substantia nigra, while enalapril treatment resulted in high cAMP levels in the ventrolateral medulla oblongata (A1 catecholaminergic cell group). 3. Both drugs, especially captopril, depleted cAMP content in the cingulate cortex. 4. No changes in cAMP levels were measured in the primary baroreceptor centre (nucleus of the solitary tract) following either captopril or enalapril treatment.     

RELAXATION FOLLOWING CONTRACTION IN TONICALLY CONTRACTED SMOOTH MUSCLE FROM THE BOVINE TRACHEA

Isometric tension was recorded from strips of bovine tracheal smooth muscle in which the tone had been artificially raised by agonist drugs such as histamine and carbachol. Application of exogenous acetylcholine produced a biphasic response consisting of an initial contraction followed by a more prolonged relaxation before tone was restored to normal. Atropine blocked both components of the biphasic response to exogenous acetylcholine. Tetrodotoxin blocked neither phase of the response to exogenous acetylcholine even though a similar biphasic response to electrical stimulation was severely disrupted. Application of exogenous substance P produced a biphasic response of similar magnitude and form to that produced by acetylcholine. Application of exogenous histamine (tone raised by carbachol) also produced a biphasic response although higher concentrations were required to produce a relaxation of equal magnitude to that produced by acetylcholine. It is concluded that the inhibitory component of the biphasic response to exogenous acetylcholine occurs as a non-specific sequel to contraction.