Milan Hypertensive Rats Research Articles

No persistent effect of angiotensin converting enzyme inhibitor treatment in Milan hypertensive rats despite regression of vascular structure.

Milan hypertensive rats were treated, from ages 4 to 24 weeks, with the angiotensin converting enzyme (ACE) inhibitor, perindopril, in doses of 1.5 mg/kg per day or 0.4 mg/kg per day. Controls were untreated Milan hypertensive rats. At age 24 weeks, a mesenteric biopsy was taken, from which two resistance vessels were taken out and mounted on a myograph for structural and functional analysis. Thereafter, treatment was withdrawn and the blood pressure of the rats was followed until age 36 weeks. Perindopril treatment had a dose-dependent effect on blood pressure as well as on both structural (media thickness, media: lumen ratio) and functional (estimated pressure against which vessels could contract) parameters of the resistance vessels. However, when treatment was withdrawn, blood pressure rose to (low-dose perindopril group) or above (high-dose perindopril group) control levels. The results contrast with previous studies using spontaneous hypertensive rats (SHR) where, using the same protocol, blood pressure remains low after withdrawal of treatment with ACE inhibitors, including perindopril, although the effect on resistance vessel parameters is similar. The results show that the persistent effect of ACE inhibitor therapy which has been seen in SHR is not a general feature of genetic hypertension. Furthermore, these results raise doubts as to whether the persistent effect seen in SHR is due to a general effect of ACE inhibitor treatment on vascular structure.

Pathogenesis of the essential hypertensions

The pathogenesis of essential hypertension (EH) is reviewed with a special focus on the development phase or the pre-hypertensive period. Three animal models are presented: the spontaneously hypertensive rat, the Dahl's salt-sensitive rat, and the Milan hypertensive rat. Some of the findings in animal models have inspired new fields and technical approaches for studying EH in man. From the original idea of Page, a new mosaic of various etiological parameters serves as a basis for reviewing the multiple facets of EH in man. One must conclude that EH is heterogeneous disease and most likely every single hypertensive patient belongs to a subgroup of the whole population of hypertensives.

Calpastatin level in spontaneously hypertensive rats

We studied calpastatin activity in erythrocytes of Milan hypertensive and prehypertensive rats, in their normotensive controls, in F1 and F2 hybrids, and in two inbred strains derived from F2, one hypertensive and the other normotensive. Our results show that the decrease in calpastatin activity observed in Milan hypertensive rats was not caused by hypertension, it was transmitted in a recessive way in heterozygous, and it was not correlated to hypertension.

49. The hypothalamic ouabain-like factor in the Milan hypertensive rats

49. The hypothalamic ouabain-like factor in the Milan hypertensive rats

Analysis of the renin gene intron A tandem repeat region of Milan and Lyon hypertensive rat strains

The region of intron A of the rat renin gene containing a unique tandemly repeated sequence was analysed in the Milan and Lyon hypertensive rat strains and their controls, and in several Sprague-Dawley rats, using an oligonucleotide probe complementary to the tandemly repeated sequence and a renin complementary DNA probe. In the Milan rats, the size of the Bgl II DNA fragment encompassing the tandem repeat region was the same in the hypertensive (MHS) and normotensive (MNS) strains. In the Lyon model, a difference of 1.1 kilobase (equivalent to about 28 copies of the 38 basepair tandem repeat sequence) was observed in the size of the Bgl II fragment of the hypertensive (LH) and normotensive (LN) strains. However, the finding that the size of the fragment in the Lyon low-blood-pressure (LL) strain was the same as that in the LH strain rather than the LN strain suggests that the difference between the two latter strains is not by itself a major cause of the blood pressure difference between them in the intron A tandem region. An analysis of Sprague-Dawley rats, from which the Lyon strains are derived, showed that at least three different renin gene alleles, two with Bgl II fragments of the same size as those seen in the Lyon strains, are randomly segregating in this population.

Vascular smooth muscle cells from the Milan hypertensive rat exhibit decreased functional angiotensin II receptors.

The present study was designed to study the functional properties of Angiotensin II (Ang II) binding sites in vascular smooth muscle cells in the Milan hypertensive rat (MHS), a model of low renin hypertension. Smooth muscle cells from MHS rats exhibited increased growth in culture in comparison with the Milan normotensive strain (MNS) as determined by population doubling times (24.5 +/- 2 and 34.8 +/- 2 hours, n = 4, respectively). Hormone receptor number, evaluated by binding assays using [125I]Ang II, showed no difference in either receptor number or affinity for both cell types. The functional responsiveness of Ang II receptors was evaluated by measuring the activation of phospholipase C, Na(+)-H+ exchange, and cytosolic Ca2+ levels. Phospholipase C activity was determined as tritium-labeled inositol trisphosphate and bisphosphate release before and after 15-second exposure to 10(-7) M Ang II. Ang II-stimulated phospholipase C activity in MNS (p less than 0.02) but not in MHS cells. Na(+)-H+ exchange was measured as the dimethylamiloride-sensitive 22Na+ influx into acid-loaded vascular smooth muscle cells with and without 10(-7) M Ang II. In MNS cells, Ang II significantly stimulated (p less than 0.001) antiporter activity but not in MHS cells, which showed a uniformly blunted response. MHS cells exhibited higher basal cytosolic Ca2+ levels than MNS cells, but Ca2+ rapidly increased in the presence of Ang II in MNS but not in MHS cells. Direct activation of phospholipase C by GTP-gamma-S in permeabilized cells indicated that both strains exhibited similar coupling levels by guanine-nucleotide binding proteins.(ABSTRACT TRUNCATED AT 250 WORDS)

Diuretic effect of bumetanide in isolated perfused kidneys of Milan hypertensive rats

The Milan hypertensive strain of rats (MHS) is characterized in the prehypertensive phase (4 weeks of age) by a significantly faster bumetanide-sensitive cell membrane Na+,K+,Cl(-)-cotransport as compared to matched normotensive controls (MNS). The isolated kidney preparation, which allows the study of renal function under controlled in vitro conditions, was chosen to compare the natriuretic effect of the loop diuretic, bumetanide, to that of two other diuretics (amiloride and hydrocholorothiazide) acting on different parts of the nephron. Concentrations ranging from 10(-7) to 10(-4) M were tested in 4-week-old MHS and MNS. Our results showed that the natriuretic response to all diuretics was greater in MHS as compared to MNS when evaluated as absolute Na+ excretion (UNA+); this is likely because of the faster basal glomerular filtration rate (GFR) in the hypertensive strain (874 +/- 126 in MHS vs. 556 +/- 33 microliters.min-1.g-1 k wt in MNS, P less than 0.05). However, when calculated either as a difference from basal values (delta UNa+), or per ml of glomerular filtration rate, the response of MHS kidneys to amiloride and hydrochlorothiazide was similar in the two strains: delta UNa+ after amiloride at (10(-4) M was + 2.1 +/- 0.7 in MHS versus + 1.2 +/- 0.2 mumol.g-1.g-1 k wt in MNS; after hydrochlorothiazide 10(-4) M it was + 1.7 +/- 0.9 in MHS versus + 1.1 +/- 0.4 mumol.min-1.g-1 k wt in MNS, values not statistically different.(ABSTRACT TRUNCATED AT 250 WORDS)

On the pathogenetic mechanism of hypercalciuria in genetically hypertensive rats of the Milan strain.

The aim of this study was to investigate the pathogenesis of hypercalciuria in the Milan strain of genetically hypertensive rats. Dietary calcium intake and urinary and fecal calcium output were measured simultaneously with indices of sodium and phosphate homeostasis in male rats of the Milan hypertensive and normotensive strains. In addition, urinary calcium and creatinine excretion rates, calcium, phosphate and creatinine serum concentrations, and bone calcium content were also measured in these rats after an overnight fast. Under fed steady-state conditions dietary calcium, sodium, and phosphate intakes, were similar in the two groups of rats, but hypertensive rats had twofold higher urinary calcium excretion and normal urinary excretion of sodium and phosphate. Fecal calcium output was slightly but significantly higher in the adult hypertensive rats while fecal sodium and phosphate excretion was normal. Because of increased urinary and fecal calcium loss, net calcium balance was significantly less positive in hypertensive than in control rats. Under fasting conditions hypertensive rats were confirmed to have hypercalciuria despite normal serum calcium concentrations and normal creatinine clearance. In accordance with balance data and fasting hypercalciuria, bone calcium content was found to be significantly reduced in hypertensive rats. These findings confirm that hypercalciuria in the Milan hypertensive rats is explained by an altered renal calcium handling; it is also associated with a slightly increased fecal calcium output and, therefore, with a less positive calcium balance and reduced bone calcium content.

Na+-H+ exchange and other ion-transport systems in erythrocytes of essential hypertensives and spontaneously hypertensive rats: a comparative analysis

The activity of ion-transport systems and Ca2+-induced erythrocyte haemolysis were compared between patients with essential hypertension and two strains of spontaneously hypertensive rats. Previous data on the increased rate of Na+-Li+ countertransport in erythrocytes of essential hypertensives were confirmed in this study. However, identification of Na+-Li+ countertransport in rat erythrocytes remained a complicated person because of the high rate of sodium-independent efflux of Li+. The rate of Na+-H+ exchange increased by 50-80% both in spontaneously hypertensive Wistar-Kyoto rats (SHR) and in patients with essential hypertension. No difference between Milan hypertensive strain rats (MHS) and Milan normotensive strain rats (MNS) was found. The rate of Na+,K+ cotransport increased in SHR and MHS erythrocytes compared with rats of the control strains [normotensive Wistar-Kyoto rats (WKY) and MNS; 30-50 and 90-110%, respectively]. No difference in this parameter was found between patients with essential hypertension and healthy subjects. Erythrocytes of patients with essential hypertension and of SHR were characterized by a higher sensitivity of their K+ channels to the increased concentration of intracellular Ca2+. This parameter did not change in MHS erythrocytes. Ca2+-induced haemolysis increased four- to fivefold in MHS erythrocytes compared with MNS and did not change in erythrocytes of SHR and patients with essential hypertension. The conclusion from these data is that the SHR strain is a more adequate model of human essential hypertension than the MHS.

Na,K-ATPase Activity in Renal Tubule Cells From Milan Hypertensive Rats

Several abnormalities of cation transport have been described in the Milan hypertensive rats (MHS). In this study we examined Na,K-ATPase activity in proximal convoluted tubules (PCT) cells and medullary thick ascending limb of Henle cells (TAL) from MHS and from the Milan normotensive rats (MNS). Na,K-ATPase activity was determined as 32P-ATP hydrolysis in single tubule segments. Na,K-ATPase activity (pmol Pi/mm t/h) was significantly higher in MHS than MNS both in PCT (903 +/- 227 n = 8 v 506 +/- 285 n = 12) and TAL (4324 +/- 800 n = 5 v 3063 +/- 625 n = 5). Na,K-ATPase dependent respiration was determined in PCT cell from MNS and MHS. Under basal condition Na,K-ATPase dependent respiration (mumol O2/mg protein/h) was higher in MHS than in MNS (24.2 +/- 1.8 n = 5 v 16.1 +/- 0.4 n = 5). When the cells were Na loaded by amphotericin Na,K-ATPase dependent respiration increased significantly more in MHS than MNS (38.4 +/- 1.6 v 26.8 +/- 2.2 n = 4). Thus, Na,K-ATPase activity is higher in renal tubule cells both at normal intracellular Na and after the cells have been Na loaded. The results indicate that regulation of Na homeostasis in renal tubule cell is different in MHS and MNS.

34 The role of membrane skeleton in the alteration of Na-K cotransport in Milan hypertensive rats

34 The role of membrane skeleton in the alteration of Na-K cotransport in Milan hypertensive rats

Calcium transport in basolateral plasma membranes from kidney cortex of Milan hypertensive rats

Ca 2+ transport was investigated in basolateral plasma membranes (BLM) isolated from kidney cortex of the Milan strain of genetically hypertensive rats (MHS) and their normotensive controls (MNS) during a pre-hypertensive stage (age 3–4 weeks). It was found that the V max of ATP-dependent Ca 2+ transport (in the presence of calmodulin) was about 16% lower in MHS than in control rats. In membranes from MNS rats which had been isolated in the presence of EGTA, the ATP-dependent Ca 2+ transport showed a hyperbolic Ca 2+ concentration dependence, a high K m (Ca 2+) and a low V max ; upon addition of exogenous calmodulin, the kinetics became sigmoidal, the K m (Ca 2+ was decreased and the V max was increased. In membranes from MHS rats, the Ca 2+ concentration dependence of ATP-driven Ca 2+ transport was sigmoidal and the Ca 2+ affinity was high in the absence of added calmodulin. Addition of exogenous calmodulin to these membranes resulted in an increase in V max , but no change in other kinetic parametrs. Low-affinity hyperbolic kinetics of Ca 2+ transport could only be obtained in MHS rats if the membranes were extracted with hypotonic EDTA and hypertonic KCl. These data suggest that the plasma membrane Ca 2+-ATPase, which catalyses the ATP-dependent Ca 2+ transport, exists in BLM of pre-hypertensive MHS rats predominantly in an activated, high-affinity form.

Ouabainlike Na+,K+-ATPase inhibitor in the plasma of normotensive and hypertensive humans and rats.

Acute volume expansion, increased sodium intake, and restraint on sodium excretion endow the plasma with an increased capacity to inhibit sodium transport. Cytochemical techniques can detect the presence of Na+K+-adenosine triphosphatase (ATPase) inhibitor in the plasma of normal humans and rats, the concentration of which is controlled by salt intake. The substance responsible appears to originate in the hypothalamus, where the concentration is also controlled by salt intake. The plasma concentration of the cytochemically detectable Na+,K+-ATPase inhibitor is substantially raised in the plasma of patients with essential hypertension, spontaneously hypertensive rats (SHR) and of Milan hypertensive rats. The concentration of activity in the hypothalamus of SHR is also considerably raised. These findings demonstrate that these forms of hypertension are associated with a rise in the concentration of a cytochemically detectable circulating Na+,K+-ATPase inhibitor that under normal circumstances is controlled by salt intake.

Decreased level of calpain inhibitor activity in kidney from Milan hypertensive rats

Rat kidney contains two different calpain isozymes distinguishable on the basis of their Ca 2+ requirement and of their activation mechanisms. The two calpain isozymes are present in comparable amounts in kidney of normotensive and hypertensive rats of the Milan strain. Conversely, the level of the natural inhibitor of calpain is significantly decreased in kidney of hypertensive rats as compared to control normotensive rats. This deficiency is more pronounced in the cortical region than in other kidney fractions. These results taken together with previous observations indicating the existence of an identical defect in red cells from the same hypertensive rat strain, (Pontremoli, S., Melloni, E., Salamino, F., Sparatore, B., Viotti, P., Michetti, M., Duzzi, L., and Bianchi, G. (1986) Biochem. Biophys. Res. Commun. 138, 1370–1375) emphasize the possible role of an unbalanced intracellular proteolytic system in the development of genetically determined hypertension.

Cytochemically assayable Na+,K+-ATPase inhibition by Milan hypertensive rat plasma.

The ability of plasma from 3- and 9-week-old Milan hypertensive rats and their normotensive controls to inhibit Na+,K+-adenosine triphosphatase (ATPase) was studied using cytochemical bioassay techniques in fresh tissue. With a validated cytochemical bioassay that measures the capacity of biological samples to stimulate glucose-6-phosphate dehydrogenase activity in guinea pig proximal tubules as an indication of their capacity to inhibit Na+,K+-ATPase, the mean glucose-6-phosphate dehydrogenase-stimulating ability of the plasma of the 9-week-old Milan hypertensive rats and their normotensive controls was 586.0 +/- 88 and 23.4 +/- 8.3 U/ml (n = 7; p less than 0.001), while that of the 3-week-old Milan hypertensive rats (before the main rise in arterial pressure) and their normotensive controls was 99.9 +/- 27.4 and 7.8 +/- 1.8 U/ml (n = 7; p less than 0.001). With the use of a semiquantitative cytochemical assay that measures Na+,K+-ATPase activity directly, plasma from the adult hypertensive rats had a much greater capacity to inhibit Na+,K+-ATPase than the plasma of the control rats. The significantly raised levels found in the young hypertensive rats before the main rise in arterial pressure are consistent with the hypothesis that the rise in the ability of plasma to inhibit Na+,K+-ATPase is due to an inherited renal difficulty in excreting sodium.

Relationship between erythrocyte volume and sodium transport in the Milan hypertensive rat and age-dependent changes.

The relationship between differences in red blood cell (RBC) volume and ion transport across the erythrocyte cell membrane were investigated in the Milan Hypertensive (MHS) and Milan Normotensive (MNS) rat strains, under different experimental conditions and during ageing. The results obtained indicate that: the difference in Na+/K+ cotransport between MHS and MNS disappear when the RBC volume of the two strains becomes equal under hypotonic swelling; MHS RBCs are osmotically more fragile than those of MNS, probably because of a different membrane structure rather than a different amount of membrane surface, and the smaller volume and the lower Na+ content of MHS RBCs are maintained throughout the life span, while Na+/K+ pump activity and Na+/K+ cotransport undergo age-dependent changes, related to the development of hypertension. All these findings suggest that a primary abnormality of the cell membrane structure of MHS, probably located in the cytoskeleton, is responsible for the cell functional alterations that we previously demonstrated to be genetically associated with MHS hypertension.

Sodium Transport in Membrane Vesicles from Kidney of the Milan Hypertensive Rat Strain

Sodium Transport in Membrane Vesicles from Kidney of the Milan Hypertensive Rat Strain

Volumes and Na Transports in Intact Red Blood Cells, Resealed Ghosts, and Inside-Out Vesicles of Milan Hypertensive Rats

Volumes and Na Transports in Intact Red Blood Cells, Resealed Ghosts, and Inside-Out Vesicles of Milan Hypertensive Rats

Sodium Transport in Membrane Vesicles from Kidney of the Milan Hypertensive Rat Strain

Sodium Transport in Membrane Vesicles from Kidney of the Milan Hypertensive Rat Strain

Volumes and Na Transports in Intact Red Blood Cells, Resealed Ghosts, and Inside-Out Vesicles of Milan Hypertensive Rats

Volumes and Na Transports in Intact Red Blood Cells, Resealed Ghosts, and Inside-Out Vesicles of Milan Hypertensive Rats