Different ratio of myosin heavy chain isoforms in arterial smooth muscle of spontaneously hypertensive rats

Summary The relative proportion of the two putative heavy chains of smooth muscle myosin (MHC1 and MHC2) was determined in the caudal and femoral arteries of spontaneously hypertensive rats (SHR) and normotensive (WKY) rats at 16 weeks of age. The heavy chain polypeptides with M_r 204000 and 200000 were resolved electrophoretically under denaturing conditions in porous polyacrylamide gels. Both proteins reacted strongly with a monoclonal antibody (2C4) to smooth muscle MHC. In caudal arteries the ratio of MHC1/MHC2 was 3.11 in SHR rats compared with 1.81 in WKY rats (p<0.005) and similarly in femoral arteries, 2.81 vs 1.51 (p<0.001). In the portal vein there was no significant difference, 1.71 vs 1.51. The possibility that the higher MHC ratio in the SHR is the genetically mediated defect in arterial smooth muscle cells leading to the hypertension is discussed as an alternative to the elevated systemic blood pressure causing the altered MHC ratio.     

Potassium relaxation of vascular smooth muscle from spontaneously hypertensive rats.

Helical strips of tail artery from spontaneously hypertensive (SHR) and Kyoto Wistar normotensive rats (WKY) were observed to relax in response to potassium after contraction induced by norepinephrine in potassium-free solution. Helical strips from SHR consistently showed greater relaxation in response to the addition of potassium than did those from WKY. The amplitude of potassium-induced relaxation is believed to be an index of the activity of electrogenic sodium-potassium transport and hence of sodium-potassium ATPase. Thus, the sodium-potassium ATPase activity of SHR vascular smooth muscle is increased as compared to WKY. This interpretation is supported by the observation that ouabain eliminated potassium-induced relaxation in both SHR and WKY strips. Potassium-induced relaxation in SHR was more sensitive to ouabain inhibition than in WKY. Relaxation induced by potassium in SHR and WKY strips was also shown to vary with: (1) the length of incubation in potassium-free solution, and (2) the concentration of potassium added back. The results of these experiments on potassium-induced relaxation serve as evidence that SHR have either an increased intrinsic sodium-potassium ATPase activity, or an enzyme activity that has been stimulated to a greater degree by an elevated intracellular sodium concentration which resulted from the incubation in potassium-free solution.     

Beta-adrenergic receptor differences in cultured arterial smooth muscle cells between spontaneously hypertensive and Wistar-Kyoto rats

In this study, beta-adrenergic receptors of cultured arterial smooth muscle cells (ASMC) from normotensive Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR) were compared. Scatchard analysis of [3H]-dihydroalprenolol hydrochloride (DHA) binding revealed that the maximum binding capacity (Bmax) for [3H]-DHA binding was significantly higher in WKY than in SHR, by twofold (n = 6, P less than 0.0001). Analysis of isoproterenol competition for [3H]-DHA binding by computerized non-linear regression indicated that the two strains have an equal number of high-affinity-state receptors, but that WKY rats have about three times more low-affinity-state receptors than SHR. Isoproterenol stimulated cyclic adenosine monophosphate (cAMP) production in a dose-dependent manner, and dose-response experiments revealed an effective concentration of ligand at which 50% of the maximum effect is observed (EC50) of 5-10 x 10(-8) mol/l isoproterenol for both WKY and SHR. Also, isoproterenol-stimulated cAMP production was inhibited by propranolol in a dose-dependent manner. This study demonstrates that the previously described difference observed for in vivo binding of beta-adrenergic receptors of vessels from SHR is retained in tissue culture of vascular smooth muscle and is, therefore, independent of the elevated plasma catecholamine and blood pressure levels found in SHR. The finding that there is equal cAMP stimulation in WKY and SHR is consistent with the discovery of an equal number of high-affinity-state receptors in the two strains. The difference in total beta-adrenergic receptor number is secondary to the greater number of low-affinity-state receptors found in WKY.     

Phytoestrogens attenuate oxidative DNA damage in vascular smooth muscle cells from stroke-prone spontaneously hypertensive rats

OBJECTIVES: A recent study demonstrated that reactive oxygen species (ROS) were involved in the maintenance of hypertension in stroke-prone spontaneously hypertensive rats (SHRSP). However, the role of oxidative stress in hypertension and its related diseases in SHRSP remains unknown. To determine whether phytoestrogens attenuate oxidative DNA damage in vascular smooth muscle cells (VSMC) from SHRSP and Wistar-Kyoto (WKY) rats, we investigated the effect of daidzein, genistein and resveratrol on oxidative DNA damage in VSMC, induced by advanced glycation end-products (AGEs). METHODS: VSMC were treated with AGEs in the presence or absence of phytoestrogens for the indicated time. Cellular degeneration induced by AGEs was characterized in terms of intracellular oxidant levels, intracellular total glutathione (GSH) levels, mRNA expression for gamma-glutamylcysteine synthetase (GCS), and a new marker of oxidative stress, 8-hydroxy-2'-deoxyguanosine (8-OHdG) contents. RESULTS: AGEs stimulated 8-OHdG formation in VSMC in a time- and dose-dependent manner. We also confirmed that VSMC from SHRSP were more vulnerable to oxidative stress induced by AGEs, than VSMC from WKY rats. Daidzein, genistein or resveratrol reduced AGEs-induced 8-OHdG formation in a dose-dependent manner. The preventive effects of phytoestrogens on 8-OHdG formation remarkably paralleled changes in the intracellular oxidant levels in VSMC following AGEs treatment. We further demonstrated that phytoestrogens increase intracellular total GSH level in VSMC. Increased GSH synthesis was due to enhanced expression of the rate-limiting enzyme for GSH synthesis, GCS. Phytoestrogens-stimulated total GSH level in VSMC could lead to decreased intracellular oxidant levels, and thus prevent oxidative DNA damage, induced by AGEs. The phytoestrogens are powerful antioxidants able to interfere with AGEs-mediated oxidative DNA damage of VSMC, and are potentially useful against vascular diseases where ROS are involved in hypertension.     

RhoA activation in vascular smooth muscle cells from stroke-prone spontaneously hypertensive rats.

RhoA is commonly activated in the aorta in various hypertensive models, indicating that RhoA seems to be a molecular switch in hypertension. The molecular mechanisms for RhoA activation in stroke-prone spontaneously hypertensive rats (SHRSP) were here investigated using cultured aortic smooth muscle cells (VSMC). The level of the active form of RhoA was higher in VSMC from SHRSP than in those from Wistar-Kyoto rats (WKY). The phosphorylation level of myosin phosphatase target subunit 1 (MYPT1) at the inhibitory site was also significantly higher in SHRSP, and the phosphorylation levels in both VSMCs were strongly inhibited to a similar extent by treatment with Y-27632, a Rho-kinase inhibitor. The expression levels of RhoA/Rho-kinase related molecules, namely RhoA, Rho-kinase, MYPT1, CPI-17 (inhibitory phosphoprotein for myosin phosphatase) and myosin light chain kinase, were not different between SHRSP and WKY. Valsartan, an angiotensin II (Ang II)- type 1 receptor antagonist, selectively and significantly reduced the RhoA activation in VSMC from SHRSP. The expression levels of the Rho GDP-dissociation inhibitor (RhoGDI) and leukemia-associated Rho-specific guanine nucleotide exchange factor (RhoGEF) did not differ between SHRSP and WKY. In cyclic nucleotide signaling, cyclic GMP (cGMP)-dependent protein kinase Ialpha (cGKIalpha) was significantly downregulated in SHRSP cells, although there were no changes in the expression levels of guanylate cyclase beta and cyclic AMP (cAMP)-dependent protein kinase or the intracellular contents of cGMP and cAMP between the two rat models. These results suggest that the possible mechanisms underlying RhoA activation in VSMC from SHRSP are autocrine/paracrine regulation by Ang II and/or cGKIalpha downregulation.     

Arterial smooth muscle contractions in spontaneously hypertensive rats on a high-calcium diet.

OBJECTIVE: To study the effects of a high-calcium diet upon blood pressure, vascular smooth muscle contractions and intracellular free calcium in spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto (WKY) rats. DESIGN: Eight-week old animals were placed on a normal-calcium diet (1.1% calcium; SHR and WKY rat groups) or a high-calcium diet (2.1% calcium; Ca-SHR and Ca-WKY rat groups) and observed for 12 weeks. METHODS: Blood pressure was measured indirectly by the tail-cuff method and in vitro smooth muscle responses were studied using a standard organ bath chamber. Platelets were used as a cell model for analysis of intracellular free calcium concentration, measured by the fluorescent indicator Quin-2. RESULTS: The blood pressure of Ca-WKY and WKY rats did not differ, but increased systolic blood pressure was attenuated in Ca-SHR compared with SHR. The concentration-response curves of mesenteric arterial rings for potassium chloride and noradrenaline were not affected by the high-calcium diet in either SHR or WKY rats. The time required for total relaxation after washout of contractile agents (washout time) was shortest in WKY and Ca-WKY rats after both agonists, and shorter in Ca-SHR than in SHR after noradrenaline. Smooth muscle responses were also studied by contracting the preparations with noradrenaline and potassium chloride in a calcium-free solution, after which, calcium was added to the organ bath in increasing concentrations. Calcium contraction responses were similar in WKY and Ca-WKY rats; SHR displayed an attenuated response to calcium addition in mesenteric rings stimulated by both agonists. After potassium chloride as agonist, the responses of SHR and Ca-SHR did not deviate but, after noradrenaline, a significant shift in the calcium contraction curve towards the normotensive curve was observed in Ca-SHR. Intracellular free calcium was clearly lower in WKY rats than in SHR, and was significantly reduced by calcium supplementation in the hypertensive but not the normotensive animals. CONCLUSIONS: A reduction in intracellular free calcium concentration and an effect upon receptor-mediated vascular smooth muscle contraction and excitation-contraction coupling may participate in the blood pressure lowering effect of a high-calcium diet.     

The contractility of venous vascular smooth muscle in spontaneously hypertensive or renal hypertensive rats

Summary The influence of arterial hypertension on the contractility of venous smooth muscle was studied by using force-velocity relations derived from the isolated tetanized portal vein of spontaneously hypertensive (SHR) or renal hypertensive rats (RHR). The results were compared with those obtained from corresponding normotensive rats (NR).The portal vein contractility of SHR was determined by the analysis of either a single isometric contraction or of a number of afterloaded isotonic contractions. The series elasticity needed to calculate the shortening velocity of the contractile element was found to be greater in SHR than in NR. Both the isometric and the isotonic force-velocity relations were shifted similarly by the spontaneous hypertension. The maximum rate in tension increase was greater by a factor of 1.4 and the force generation was increased by a factor of 1.5 as compared with the results obtained from the NR. The velocity of shortening of the unloaded preparation remained constant, which indicates that the turnover rate of the myosin linkages, i.e. the elementary process of contraction, was unchanged (the so-called polytropic effect). The augmentation in force generation is probably caused by a recruitment process induced by an increase in the intracellular calcium level.In RHR a slight increase in the force generation of the portal vein from 15.6±1.1 mN to 18.4±1.0 mN was seen in rats which had been submitted to a period of arterial hypertension of about 20 weeks. The speed of isotonic shortening extrapolated to zero load was considerably reduced from 0.86±0.03 ML/s to 0.61±0.09 ML/s (P<0.0025); this is the so-called tachytropic effect.The results revealed a different influence on the elementary process of vascular smooth muscle contraction for both types of arterial hypertension. The dynamics of the cycling myosin linkages were only affected by renal hypertension whereas with spontaneous hypertension the dynamics remained constant.

---

Die Kontraktilität venöser Gefäßmuskeln von spontan hypertensiven oder renal hypertensiven Ratten

Zusammenfassung Der Einfluß arterieller Hypertonie auf die Kontraktilität von Gefäßmuskeln wurde über Kraft-Geschwindigkeits-Beziehungen an der isolierten, tetanisierten Portalvene spontan-hypertensiver (SHR) und renal-hypertensiver Ratten (RHR) untersucht.Bei spontan-hypertensiven Tieren (n=23, mittlerer arterieller Blutdruck=166±4 mm Hg) wurden die Kraft-Geschwindigkeits-Beziehungen aus einer größeren Anzahl nachbelasteter isotoner Kontraktionen oder aus einzelnen isometrischen Kontraktionen unter Berücksichtigung der Serienelastizität berechnet. Die Serienelastizität war bei den SHR-Tieren größer als bei der Vergleichsgruppe normotoner Tiere (NR). Sowohl die isometrische als auch die isotonische Kraft-Geschwindigkeits-Beziehungen wurden durch die spontane Hypertonie in gleicher Weise beeinflußt. Verglichen mit den Ergebnissen an NR nahm die maximale Geschwindigkeit des Kraftanstiegs um den Faktor 1.4 zu. Die Kraftamplitude stieg um den Faktor 1.5 an, während die Geschwindigkeit der lastfreien Verkürzung als ein Maß für die Geschwindigkeit der Querbrückenbewegung konstant blieb (sog. polytroper Effekt).Die Veränderung der Kontraktilität der Portalvene in SHR ist somit nicht durch eine Beeinflussung der Dynamik der einzelnen Querbrücken hervorgerufen worden; der elementare Kontraktionsprozess bleibt unbeeinflußt. Die gesteigerte Kraftentwicklung ist möglicherweise auf einen durch Anstieg des intrazellulären Kalzium-Spiegels bedingten Rekrutierungsprozeß zurückzuführen.Bei den renal hypertensiven Tieren (Gruppe I: 75 Tage nach Klammerung der linken Nierenarterie, n=14, mittlerer arterieller Blutdruck=175±2 mm Hg; Gruppe II: 138 Tage nach der Klammerung, n=4, 177±10 mm Hg) war ein geringer Anstieg der Kraftentwicklung der Portalvene vor allem in Untersuchungen an Ratten mit langdauernder Hypertonie (RHR II) zu beobachten (15.6±1.1 mN in NR, 18.4±1.0 mN in RHR II). Die lastfreie Verkürzungsgeschwindigkeit wurde deutlich von 0.86±0.03 ML/s auf 0.61±0.09 ML/s vermindert (P<0.0025; sog. tachytroper Effekt). Die erhobenen Befunde lassen einen unterschiedlichen Einfluß der beiden Formen einer arteriellen Hypertonie auf den elementaren Kontraktionsprozeß der Portalvene der Ratte erkennen. Die Dynamik des Querbrückenzyklus wird nur bei der renalen Hypertonie, nicht aber bei der spontanen Hypertonie beeinflußt.

---

---

With 4 figures and 3 tables     

Alterations of calcium channels in vascular smooth muscle cells from spontaneously hypertensive rats.

We examined the possible alterations in calcium handling through the calcium channels of spontaneously hypertensive rats (SHR) using 45Ca2+ uptake measurements in cultured aortic cells. Primary cultures of vascular smooth muscle cells (VSMC) were obtained by enzymatic dissociation of the thoracic aortas from 8-week-old SHR and age-matched Wistar-Kyoto rats (WKY). The functions of voltage sensitive calcium channels (VSCC) and receptor operated calcium channels (ROCC) were estimated from the activated 45Ca2+ uptake in VSMC with high K+ depolarization and arginine vasopressin (AVP), respectively. Compared to basal conditions, depolarization with 55 mM KCl increased 45Ca2+ uptake at 20 min by 94 +/- 17 (SE) % in SHR and 38 +/- 6% in WKY. The activated 45Ca2+ uptake was significantly greater in SHR than in WKY (p < 0.01). There was no significant difference in 45Ca2+ uptake at 20 min in the presence of 5 x 10(-8)M AVP between SHR and WKY. These results suggest that calcium uptake, at least through VSCC, is increased in VSMC of SHR. This enhanced activity may be implicated in the hypertensive mechanisms in this model of hypertension.     

Long-term arterial pressure in spontaneously hypertensive rats is set by the kidney.

OBJECTIVES: We investigated whether arterial pressure in spontaneously hypertensive rats (SHR) can be normalized by a kidney graft from normotensive histocompatible donors. In addition, the effect of differential genetic predisposition to hypertension of recipients of an SHR kidney on the development of post-transplantation hypertension was studied. METHODS: SHR were transplanted with a kidney from congenic rats (BB.1K) homozygous for a 2 cM segment of SHR chromosome 20, including the major histocompatibility complex class Ia and class II genes. BB.1K and F1 hybrids (F1H, SHR x Wistar-Kyoto rats) were transplanted with an SHR kidney and the development of renal post-transplantation hypertension was monitored. RESULTS: Thirty days after renal transplantation, mean arterial pressure (MAP) was 116 +/- 4 mmHg in SHR with a BB.1K kidney (n = 8) versus 168 +/- 2 mmHg in sham-operated SHR (n = 10); P < 0.001. Cumulative renal sodium balance (mmol/100 g body weight) over 21 days after bilateral nephrectomy was 6.8 +/- 0.6 in SHR with a BB.1K kidney versus 10.8 +/- 1.6 in sham-operated SHR (P < 0.05). Within 60 days of transplantation, MAP increased in BB.1K and in F1H transplanted with an SHR kidney (n = 7 per group) by 38 +/- 5 mmHg and 43 +/- 8 mmHg, respectively. CONCLUSIONS: In SHR, arterial pressure can be normalized by a kidney graft from normotensive donors. The genetic predisposition of the recipients to hypertension does not modify the rate and the extent of the arterial pressure rise induced by an SHR kidney graft.     

Evidence for increased in vivo sodium-potassium pump activity and potassium efflux in skeletal muscle of spontaneously hypertensive rats

We have used 87Rb nuclear magnetic resonance spectroscopy (NMR) to study in vivo rubidium kinetics in spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) controls, using rubidium as a marker for potassium. We gave 15 male, 13-week-old SHR, mean +/- s.d. blood pressure 180 +/- 10 mmHg, and 15 age-matched normotensive controls, mean blood pressure 120 +/- 9 mmHg, a daily dose of RbCl (2 mmol/kg intraperitoneally). We made repeated NMR measurements of skeletal muscle rubidium concentrations until steady state was reached. We then withdrew rubidium and made further measurements of rubidium concentrations, at intervals, for up to 1 week after the last injection. We also measured plasma and erythrocyte rubidium concentrations by flame atomic absorption spectroscopy at similar intervals after the withdrawal of rubidium. Rubidium concentrations rose at a faster rate in SHR skeletal muscle, but the steady-state muscle rubidium concentration was the same (45 mmol/l) in both SHR and WKY rats. There was also a threefold increase in the rate of rubidium efflux from both muscle and erythrocytes in SHR. These results are consistent with a marked increase in Na+,K(+)-ATPase activity and an increase in the rate of rubidium efflux in vivo in SHR. The increased rate of rubidium efflux in SHR could represent increased K+ efflux via calcium-activated K+ channels and/or result as part of cell volume regulation secondary to increased Na(+)-H+ antiporter activity.     

Enhanced growth rate of cultured smooth muscle cells from spontaneously hypertensive rats

Summary Smooth muscle cells were isolated and cultured from the aortic media of age-matched, stroke-prone and stroke-resistant spontaneously hypertensive rats (SHRSP, SHRSR: SHRs) and Wistar-Kyoto rats (WKY), and the growth rate of cells from the three strains was compared. Under electron microscopical observation the cells were identified as modified (or synthetic-type) smooth muscle cells. Cells occurring at low densities showed the same morphology for both SHRs and WKY, but cells occurring at higher densities were observed to be smaller in SHRs than in WKY. An analysis of the growth curves of cells showed a significantly enhanced replication rate in cells from SHRs compared with those from WKY, especially in the early passages. In later passages (repeated until the 9th passage), however, this distinction was not clear. These growth characteristics were also confirmed in cells from both 12- and 24-week-old, age-matched SHRs and WKY. We could not find any difference between the growth characteristics of cells from SHRSP and SHRSR. It is possible to hypothesize from these findings that the abnormality relating to hypertension (hypertension being a common characteristic of SHRSR and SHRSP) is found in the smooth muscle cells, and is reflected as an enhanced growth rate when they are exposed to mitogenic stimuli, such as in atherosclerosis.     

Increased expression of osteoprotegerin in vascular smooth muscle cells from spontaneously hypertensive rats

Osteoprotegerin (OPG) is a secreted protein of the tumor necrosis factor receptor family,whichregulates bone mass by inhibiting osteoclast differentiation and activation.Although OPG is expressed ubiquitouslyand abundantly in many tissues and cell types including vascular cells,the role of OPG in other tissues is unknown.Our previous studies demonstrated that OPG was highly expressed in vascular smooth muscle cells (VSMC) andupregulated during vascular lesion formation.Methods and Results We documented,by Northern blot analysis,that the expression of OPG was more prevalent in the aorta and cultured VSMC from spontaneously hypertensive rats(SHR) compared to Wistar-Kyoto rats (WKY).In addition,we found that the expression of Angiotensin Ⅱ (Ang Ⅱ)type Ⅰ receptor (AT1R) in SHR VSMC was at significantly increased levels than in WKY VSMC.Furthermore,AngⅡ potently induced the expression of OPG in VSMC in a time- and dose-dependent manner through the AT1Rsignaling pathway.Conclusions OPG expression was substantially greater in SHR VSMC,suggesting that OPGmay be an important determinant of vascular remodeling in SHR.(J Ceriatr Cardiol 2004;1:49-54.)     

Mechanical and morphological properties of arterial resistance vessels in young and old spontaneously hypertensive rats.

We studied alterations in structural and mechanical properties of mesenteric arterial resistance vessels from young (6-week) and old (50-week) spontaneously hypertensive (SHR)and matched normotensive Wistar-Kyoto (WKY) rats. Emphasis was placed upon relating the active tension capabilities of these vessels to their smooth muscle cell content. Cylindrical segments, 0.7 mm long with internal diameters of 150 micrometer, were mounted in a myograph capable of recording circumferential vessel wall tension and dimensions. Comparisons of vessel morphology and mechanics were performed at a normalized internal circumference, L1,where active tension (delta T1) is near maximum. Arterial wall and medial hypertrophy were observed in young and old SHR. Since the percent smooth muscle cells within the media for SHR was similar to that of WKY, both increased smooth muscle cell and connective tissue content account for the medial hypertrophy. These differences in SHR vessels were reflected directly in their passive and active mechanical properties. Fully relaxed vessels from SHR were less compliant, and upon activation at L1 (high potassium depolarization), delta T1 was not different for young SHR and WKY, but values for old SHR were 35% greater (P less than 0.05) than for WKY. When relating the active force generation of the vessel to the actual smooth muscle cell area, values for smooth muscle cell stress (force/area) were similar for SHR and WKY at both ages. In addition, similarities were observed for active dynamic mechanical measurements of Young's modulus and half response time. Genetic hypertension in rats therefore appears to be associated with the development of increased vessel contractility determined by a greater number of smooth muscle cells which possess contractile properties similar to those of normotensive vessels.     

Effect of aspirin on the contractility of aortic smooth muscle in female spontaneously hypertensive rats.

The effects of acetylsalicylic acid (ASA), on aortic smooth muscle contractility during hypertension were studied in female SHR and WKY rats. The rats were administered two intraperitoneal injections of 10 mg/kg of ASA per week for three weeks. Twenty four hours after the last injection the aortic smooth muscles were evaluated for generation of active tension in response to KCl, phenylephrine, clonidine and norepinephrine. We report that aortic rings from ASA-treated SHR animals were more responsive than rings from non-treated SHR female rats. ASA treatment of SHR animals restored the contractile response to the level shown by non-treated WKY control female rats. The response from aortic rings of ASA-treated SHR to KCl, phenylephrine and clonidine was essentially similar to the response of rings from non-ASA-treated WKY control female rats. We did not observe any decrease in the systolic blood pressure during the ASA treatment in SHR female rats. These results suggest that acetylsalicylic acid modulates aortic smooth muscle contractility either through the metabolites of arachidonic acid or through alpha-adrenoceptors.     

In vivo sugar diffusion in the ileal epithelium of spontaneously hypertensive rats

BACKGROUND: D-glucose absorption, distinguishing between active and diffusive components, was studied in the ileum of spontaneously hypertensive rats (SHR) and their normotensive control Wistar-Kyoto (WKY) rats. Net water transport was also determined. METHODS: A perfusion system in vivo with ileum loops was used, and experiments with phlorizin, phloretin and 2,4,6-triaminopyrimidine (TAP) were performed in order to discriminate between active and diffusive components and between transcellular and paracellular routes. RESULTS: A significant decrease in total D-glucose absorption was found in SHR compared to WKY rats, this reduction being due to a lower SGLT1-mediated component. The effect was not compensated by the total diffusive component, since the phlorizin-insensitive D-glucose absorption did not significantly change between rat strains. However, the diffusive component of D-glucose transport was relatively more important in hypertensive than in normotensive rats. The use of 2,4,6-triaminopyrimidine (TAP), which blocks the transport across the paracellular route, showed that the paracellular diffusion of D-glucose was higher in SHR than in WKY rats. Intestinal net water absorption was not modified between either group of animals, though the presence of phlorizin in the perfusate decreased the ileal water absorption to a greater extent in normotensive rats. CONCLUSION: The observed reduction in D-glucose absorption in vivo in the ileum of SHR was due to a decrease in the SGLT1-active component. Despite the paracellular diffusion of D-glucose being higher in hypertensive than in normotensive rats, the total diffusion component was not high enough to compensate this alteration.     

Decreased responsiveness to beta-adrenoceptor agonists in arterial strips from spontaneously hypertensive rats is not associated with alterations in beta-adrenoceptors.

Beta-adrenoceptors in femoral and mesenteric arteries from 13-week-old spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto (WKY) rats were studied using radioligand binding assays and were compared with in vitro measurements of beta-adrenoceptor-mediated relaxation. The relaxant responses to noradrenaline via beta-adrenoceptors were significantly decreased in the SHR femoral artery when compared with the WKY femoral artery. However, under the same conditions, arterial relaxant responses to forskolin, an activator of adenylate cyclase, were not significantly different between SHR and WKY rats. Specific binding of 125I-iodocyanopindolol to membranes prepared from femoral arteries of SHR and WKY rats was saturable and of high affinity. Neither the equilibrium dissociation constant of 125I-iodocyanopindolol, nor the maximum number of binding sites were significantly different between SHR and WKY rats. Similar results were obtained in the case of mesenteric arteries from SHR and WKY rats. These results indicate that the decreased responsiveness to beta-adrenoceptor stimulation in SHR arteries is not associated with alterations in beta-adrenoceptors and further support the hypothesis that a reduced function of the stimulatory guanosine triphosphate-binding protein is responsible for the decreased responsiveness to a variety of receptor agonists whose mechanism of action involves adenylate cyclase activation.     

Altered calcium sequestration by subcellular fractions of vascular smooth muscle from spontaneously hypertensive rats.

The calcium sequestration characteristics of microsomal and mitochondrial fractions prepared from aortae of spontaneously hypertensive (SHR), Kyoto Wistar normotensive (NWR), and normotensive Sprague-Dawley (NSDR) rats were studied. Calcium uptake by the microsomal vesicles of SHR was significantly lower as compared to NWR and NSDR. However, mitochondria isolated from SHR aortae did not differ from NWR in their calcium uptake characteristics. Microsomal vesicles of NSDR aortae accumulated consistently more calcium than NWR. These findings illustrate the significance of relevant controls for SHR. Calcium-dependent ATPase activity in the microsomal fraction was significantly increased in SHR as compared to NWR. This may indicate a compensatory phenomenon on the part of the smooth muscle cell to increase Ca²⁺ uptake by the microsomal vesicles. These results suggest that a decreased ability to sequester calcium by the microsomal fraction of vascular smooth muscle of SHR may result in altered calcium distribution in the muscle cell.     

Neurotrophin 3 is increased in the spontaneously hypertensive rat.

OBJECTIVE: To determine whether the noradrenergic sympathetic hyperinnervation in the spontaneously hypertensive rat (SHR), a genetic model of essential hypertension, is associated with changes in neurotrophin 3 (NT3) concentrations. METHODS: NT3 levels were measured using a sensitive enzyme-linked immunosorbent assay (ELISA) in the superior cervical ganglia (SCG), heart, mesenteric artery (MA) and blood of postnatal and mature SHR and normotensive Wistar-Kyoto (WKY) rats. RESULTS AND CONCLUSIONS: NT3 levels in SHR are significantly higher in the SCG during the first 4 postnatal weeks, and in the heart and MA from 2 to 10 weeks of age, compared with levels in WKY rats. The elevated NT3 found in the sympathetic ganglia and hyperinnervated organs of SHR indicates that NT3 may play an important role in the development of hyperinnervation, possibly by enhancing the survival and/or nerve sprouting of sympathetic neurons.     

Enhanced responsiveness to angiotensin II in vascular smooth muscle cells from spontaneously hypertensive rats is not associated with alterations in protein kinase C

This study compares vascular smooth muscle cells from spontaneously hypertensive and normotensive Wistar-Kyoto rats with respect to protein kinase C and intracellular responses to angiotensin II (Ang II). Ang II-induced degradation of polyphosphoinositides and accumulation of inositol di- and tris-phosphates was enhanced (approximately twofold) in hypertensive-derived cells, without a change (vs. normotensive-derived cells) in half-maximally effective concentrations of Ang II. Intracellular pH (approximately 6.6) was comparable between both cell isolates at quiescence, but alkalinization induced by Ang II, serum, or phorbol ester was greater (delta 0.1-0.2 pH units) for hypertensive-derived cells. For both cell types, the intracellular pH response to these agonists was prevented in the presence of Na+-H+ exchange inhibitors. S6 kinase activation induced by Ang II was enhanced (approximately twofold) in hypertensive-derived cells, whereas activation in response to serum or 12-O-tetradecanoylphorbol 13-acetate did not differ significantly between the two cell types. Quantitation of protein kinase C by immunoblotting and [3H]phorbol dibutyrate binding procedures revealed no differences between the two smooth muscle cell isolates (at quiescence or in the presence of serum) with respect to either total amounts or subcellular distribution. Sensitivity of protein kinase C to phorbol ester was apparently also not different between the two cell types, as assessed from dose-dependent (phorbol ester) S6 kinase activation profiles. Phorbol ester caused a similar subcellular redistribution of [3H]phorbol dibutyrate binding in the two cell isolates, but for both, minimal (10%) translocation occurred in response to Ang II. The data suggest that enhanced agonist responsiveness in vascular smooth muscle cells is unlikely to involve alterations in protein kinase C.