Ethanol counteraction of I1-imidazoline but not alpha-2 adrenergic receptor-mediated reduction in vascular resistance in conscious spontaneously hypertensive rats

Our recent findings have shown that ethanol selectively counteracts decreases in blood pressure (BP) evoked via activation of central I1-imidazoline receptors but not alpha-2 adrenoceptors in conscious spontaneously hypertensive rats (SHRs). This study investigated the role of sympathetic activity, cardiac output and total peripheral resistance (TPR) in the differential effect of ethanol on centrally mediated hypotension. Changes in plasma norepinephrine (NE), as index of sympathetic activity, BP, heart rate, cardiac index, stroke volume, and TPR elicited by rilmenidine or alpha-methylnorepinephrine (selective I1 and alpha-2 receptor agonists, respectively) and subsequent ethanol (0.5 or 1 g/kg) or saline, were evaluated in conscious SHRs. Intracisternal rilmenidine (25 microg) or alpha-methylnorepinephrine (alpha-MNE; 4 microg) elicited similar decreases in BP, TPR, and plasma NE, but cardiac index was not changed. Ethanol (0.5 g/kg i.v.) had no effect on hemodynamic responses to rilmenidine or alpha-MNE. The higher dose (1 g/kg i.v.) of ethanol counteracted the hypotensive response to rilmenidine and significantly (P <.05) elevated TPR and plasma NE. In contrast, ethanol (1 g/kg) had no effect on the hypotensive responses to alpha-MNE but significantly (P <.05) elevated plasma NE. However, this increase in NE was approximately one third of the increase evoked by ethanol when given after rilmenidine. These findings suggest that the selective counteraction by ethanol of the hypotension evoked via activation of central I1 but not alpha-2 receptors may relate, at least in part, to its greater ability to reverse the sympathoinhibition and the associated decrease in vascular resistance mediated by I1 receptors.     

Long-term alpha 1 blockade does not reverse cardiac hypertrophy in spontaneously hypertensive rats

Not all antihypertensive drugs induce regression of left ventricular (LV) hypertrophy in hypertension, although they may equally lower blood pressure. The effects of alpha 1-blockers on regression have been inconsistent. In this study, bunazosin, a selective alpha 1-blocker, (15 mg/kg/day in food) was given to male spontaneously hypertensive rats (SHR) from 15 to 35 weeks of age to evaluate its effects on cardiac hypertrophy, hemodynamics, and neurohumoral factors. Age- and sex-matched SHR served as controls. LV function and cardiac output were determined by a micromanometer and thermodilution, respectively. Bunazosin significantly decreased blood pressure in conscious rats (from 209 to 192 mmHg, p < 0.01) but did not reduce LV mass. Heart rate, LV end-diastolic pressure, dp/dtmax, and cardiac output were similar in the 2 groups. Plasma renin activity was unaltered but plasma norepinephrine levels were higher in the treated rats (p < 0.05). Thus, bunazosin produced a significant relative reduction of blood pressure but did not reverse LV hypertrophy in SHR. Inadequate afterload reduction (8%) due to severe hypertension (> 200 mmHg) may explain the absence of regression. The rise of plasma norepinephrine levels may also offset the beneficial effects of bunazosin.     

ANP enhances bradycardic reflexes in normotensive but not spontaneously hypertensive rats

Baroreflex control of heart rate in spontaneously hypertensive rats (SHR) is defective, largely because of a poor vagal contribution to the reflex. We have demonstrated previously that atrial natriuretic peptide (ANP) enhances reflex bradycardia in normotensive rats through an action on nonarterial vagal afferent pathways. In the present study, we investigated whether ANP could reverse the baroreflex abnormality in SHR. Heart rate reflexes were activated by three different methods in conscious, instrumented SHR and Wistar-Kyoto rats (WKY) in the presence of intravenous infusions of vehicle (saline) or rat ANP (150 ng/kg per minute). Heart rate responses were measured by (1) the steady-state changes in blood pressure after alternating slow infusions (over approximately 15 to 30 seconds) of a pressor (methoxamine) and depressor (nitroprusside) drug (stimulating predominantly arterial baroreceptors), (2) the ramp method of rapid infusion of methoxamine (over < 10 seconds; stimulating arterial and cardiopulmonary baroreceptors), and (3) the von Bezold-Jarisch method of activating chemically sensitive cardiac receptors through serotonin injections. ANP enhanced the heart rate range of the arterial baroreflex (steady-state method) by 13 +/- 3% in WKY but had no significant effect on the sensitivity or any other parameter of the steady-state baroreflex. When a very rapid rise in blood pressure was elicited by the ramp method in WKY, ANP significantly enhanced baroreflex bradycardia (sensitivity increased by 29 +/- 9%, P < .05). ANP also enhanced the bradycardia of the von Bezold-Jarisch reflex (by 33 +/- 16%, P < .05) in WKY. By contrast, ANP did not influence baroreceptor or chemoreceptor heart rate reflex responses in SHR. We conclude that in normotensive rats, ANP facilitates cardiopulmonary bradycardic reflexes. The lack of effect of ANP in SHR may be related to an underlying structural or genetic alteration in their cardiac sensors, perhaps associated with cardiac hypertrophy, that prevents the ANP-induced activation of cardiac sensory afferents, resulting in cardioinhibition.     

Synergism of constitutive activity in alpha 1-adrenergic receptor activation

Recently a number of mutations have been found in vitro which maintain alpha 1-adrenergic receptors (ARs) in a partially activated form. We have previously identified two amino acid residue positions in the alpha 1b-adrenergic receptor (AR), Cys128 and Ala204, one in each of the third and fifth transmembrane segments, that constitutively activate the receptor when substituted for a phenylalanine or valine, respectively [Perez et al. (1996) Mol. Pharmacol. 49, 112-122; Hwa et al. (1996) J. Biol. Chem. 271, 7956-7964]. Another mutation analyzed previously, Ala293Glu, located in the third intracellular loop, also constitutively activates the receptor [Kjelsborg et al. (1992) J. Biol. Chem. 267, 1430-1433]. All three mutations displayed similar manifestations of constitutive activity such as higher binding affinity and potency for agonists as well as higher basal signal transduction as predicted by the revised ternary complex model of receptor activation. We hypothesized that the individual mutations because of their critical location alter the conformation of the transmembrane helices such that mimicry occur that partially conforms to the activated state, R*. To explore whether these potential conformations are independent, we combined these three mutations in all possible permutations. The combined triple mutation displays 700-fold higher binding affinity for (-)-epinephrine and 20-fold higher basal IP3 release than the wild-type receptor. We also observed that each mutation contributed independently and synergistically to both receptor agonist binding and activation with the combined mutations basal activity exceeding that of the fully-stimulated wild-type receptor. There was also a direct correlation between epinephrine's binding affinity and the degree of constitutive activity. Because the mutations affect different transmembrane domains, these results are consistent with a mechanism that helical movement acts in a concerted fashion in agonist-induced activation, a synergism predicted if multiple helix movement is involved in receptor activation.     

alpha1-Adrenoceptor subtypes mediating the contraction of arteries in spontaneously hypertensive rats

1. This study was undertaken to determine which alpha1-adrenoceptor (AR) subtypes were involved in the activation of the femoral artery and the aorta by norepinephrine (NE) in spontaneously hypertensive rats (SHR). 2. Negative log EC50 values and maximum responses of NE-induced contraction of the SHR femoral artery were unchanged and increased, respectively, compared with those of Wistar-Kyoto (WKY) rats. 3. Contractile responses of the SHR aortas to NE were similar to those of the normotensive tissues. 4. Schild plot data for alpha1-AR antagonists indicated that alpha1-AR in the aorta were predominantly alpha1H subtype. In the femoral artery, because Schild plots for bunazosin had slopes of less than 1.0, there were alpha1H (or alpha1C) and alpha1L subtypes. 5. The alpha1-AR subtype in the aorta was essentially identical to the alpha1H subtype in the femoral artery. alpha1-AR subtypes mediating contraction in the SHR blood vessels were identical to those in the WKY tissues.     

Acute mental stress but not enforced muscle activity transiently increases natural cytotoxicity in spontaneously hypertensive rats

The influence of acute mental stress and the effect of electrically induced skeletal muscle contractions on natural cytotoxicity in vivo was investigated in spontaneously hypertensive rats Natural cytotoxicity in vivo was measured as the clearance of injected 51Cr-labelled YAC-1 lymphoma cells from the lungs, which are specifically lysed by natural killer cells. The mental stress consisted of an air jet directed towards the animals in their cage for 25 min. During the mental stress there was a significant increase in natural cytotoxicity. Thus, retained radioactivity in the lungs was decreased to 74 +/- 6% of the control levels which was set to 100% (P < 0.01). This augmentation of YAC-1-cell clearance could be blocked with the beta-adrenergic receptor antagonist Timolol. Two hours after termination of the air stress, in vivo cytotoxicity had returned to control levels. In contrast, acute physical stress, consisting of electrically induced muscle contractions for 60 min, had no significant effects on in vivo cytotoxicity, either during the stimulation or 1, 2 or 24 h after the stimulation. Further, significantly increased plasma levels of adrenaline were seen after the air jet stress, but not after muscle stimulation. There were no significant changes in plasma noradrenaline levels either after air stress or muscle stimulation. These results indicate that changes in in vivo cytotoxicity after mild mental stress are dependent on increased plasma catecholamine levels while acute physical stress without changes in catecholamine levels, does not influence in vivo cytotoxicity.     

Endothelial modulation of alpha 1-adrenoceptor contractile responses in the tail artery of spontaneously hypertensive rats

1. Vascular contraction induced by phenylephrine was studied in tail artery rings from spontaneously hypertensive (SHR) and Wistar Kyoto rats (WKY) with particular focus on the role of endothelium. The influence of receptor reserve and the density of alpha 1-adrenoceptors on the possible differences observed were also analysed. 2. Phenylephrine (0.01-100 microM) induced concentration-dependent vasoconstrictions. The maximum response (alpha, P < 0.001) was greater but the pEC50 (P < 0.05) smaller in rings from SHR than from WKY rats irrespective of the presence or absence of endothelium. 3. Removal of endothelial cells resulted in a decrease of the maximum contraction with no modification in the pEC50 in arteries from both WKY and SHR. 4. The density of alpha 1-adrenoceptors (Bmax) and the dissociation constant (KD) were found to be the same for preparations from SHR and WKY rats in [3H]-prazosin binding experiments. 5. The apparent affinity (pKA) determined by the nested hyperbolic method and the operational model was similar in tail arteries from the two rat strains, irrespective of the presence or absence of endothelium. However, in endothelium-denuded rings, the pKA value was enhanced when compared with intact rings, in both SHR and WKY rats. 6. In rings from hypertensive rats, the operational parameter maximum possible effect (Em) was greater and the agonist efficacy (tau) was smaller than in rings from normotensive rats. When the endothelium was removed log tau and Em diminished in preparations from both rat strains. 7. In summary, the increased maximum responsiveness to phenylephrine in rings from SHR could be due to enhancement in Em. The log tau values indicate a deterioration in the transduction of the stimulus provided by the agonist in tail arteries from hypertensive animals. This study also suggests that the absence of endothelium modifies the alpha 1-adrenoceptor-mediated vasoconstriction probably by altering the transduction signalling mechanisms. The importance of analysing the degree of endothelium functionality when comparing results from different groups of rats is stated.     

Gbeta gamma-independent coupling of alpha2-adrenergic receptor to p21(rhoA) in preadipocytes

In preadipocytes, alpha2-adrenergic receptor (alpha2-AR) stimulation leads to a Gi/Go-dependent rearrangement of actin cytoskeleton. This is characterized by a rapid cell spreading, the formation of actin stress fibers, and the increase in tyrosyl phosphorylation of the focal adhesion kinase (pp125(FAK)). These cellular events being tightly controlled by the small GTPase p21(rhoA), the existence of a Gi/Go-dependent coupling of alpha2-AR to p21(rhoA) in preadipocytes was proposed. In alpha2AF2 preadipocytes (a cell clone derived from the 3T3F442A preadipose cell line and which stably expresses the human alpha2C10-adrenergic receptor) alpha2-adrenergic-dependent induction of cell spreading, formation of actin stress fibers, and increase in tyrosyl phosphorylation of pp125(FAK) were abolished by pretreatment of the preadipocytes with the C3 exoenzyme, a toxin which impairs p21(rhoA) activity by ADP-ribosylation. Conversely, C3 exoenzyme had no effect on the alpha2-adrenergic-dependent increase in tyrosyl phosphorylation and shift of ERK2 mitogen-activated protein kinase. alpha2-Adrenergic stimulation also led to an increase in GDP/GTP exchange on p21(rhoA), as well as to an increase in the amount of p21(rhoA) in the particulate fraction of alpha2AF2 preadipocytes. Stable transfection of alpha2AF2 preadipocytes with the COOH-terminal domain of betaARK1 (betaARK-CT) (a blocker of Gbeta gamma-action), strongly inhibited the alpha2-adrenergic-dependent increase in tyrosyl phos- phorylation and shift of ERK2, without modification of the tyrosyl phosphorylation of pp125(FAK) and spreading of preadipocytes. These results show that alpha2-adrenergic-dependent reorganization of actin cytoskeleton requires the activation of p21(rhoA) in preadipocytes. Conversely to the activation of the p21(ras)/mitogen-activated protein kinase pathway, the alpha2-adrenergic activation of p21(rhoA)-dependent pathways are independent of the beta gamma-subunits of heterotrimeric G proteins.     

3H]2-(2-Benzofuranyl)-2-imidazoline, a highly selective radioligand for I2-imidazoline receptor binding sites. Studies in rabbit kidney membranes

The purpose of this study was to delineate significant differences among mild traumatic brain-injured and early onset mild dementia patients examined using the Wechsler Memory Scale-Revised subtests. In comparison to 12 mild traumatic brain-injured patients, 11 mild dementia patients scored significantly lower on Verbal Paired Associates I and II, Visual Reproduction I and II, and Visual Paired Associates I. Raw score summaries for the Wechsler Memory Scale-Revised indicated significantly lower scores on Verbal Memory, Visual Memory, and the over-all composite. General Memory for mild dementia patients in comparison to individuals who sustained a mild traumatic brain injury. Despite equivalent scores on Attention/Concentration, the dementia group did not show significantly lowered delay in memory recall by comparison with mild closed-head injured patients. The study recommends replication with much larger sample sizes to validate the results.     

Streptozotocin-induced diabetes selectively enhances antinociception mediated by delta 1- but not delta 2-opioid receptors

We assessed the effect of diabetes on antinociception produced by intracerebroventricular injection of delta-opioid receptor agonists [D-Pen2,5]enkephalin (DPDPE) and [D-Ala2]deltorphin II. The antinociceptive effect of DPDPE (10 nmol), administered i.c.v., was significantly greater in diabetic mice than in non-diabetic mice. The antinociceptive effect of i.c.v. DPDPE was significantly reduced in both diabetic and non-diabetic mice following pretreatment with 7-benzylidenenaltrexone (BNTX), a selective delta 1-opioid receptor antagonist, but not with naltriben (NTB), a selective delta 2-opioid receptor antagonist. There were no significant differences in the antinociceptive effect of [D-Ala2]deltorphin II (3 nmol, i.c.v.) in diabetic and non-diabetic mice. Furthermore, the antinociceptive effect of i.c.v. [D-Ala2]deltorphin II was significantly reduced in both diabetic and non-diabetic mice following pretreatment with NTB, but not with BNTX. In conclusion, mice with diabetes are selectively hyper-responsive to supraspinal delta 1-opioid receptor-mediated antinociception, but are normally responsive to activation of delta 2-opioid receptors.     

Angiotensin II type 1 receptor mRNA levels in the brains of normotensive and spontaneously hypertensive rats

The type 1 angiotensin II (AII) receptor (AT1-R) has been implicated in the physiological actions mediated by AII in the brain. In view of the reported hyperactivity of the brain AII system in the spontaneously hypertensive rat (SHR), we compared the expression of AT1-R mRNAs in the brains of normotensive [Wistar Kyoto (WKY)] and SHR animals. Northern blot analysis showed about three- and approximately 20-fold increases in the levels of AT1-R mRNAs from the hypothalamus and brainstem areas, respectively, of the SHR compared with the WKY rat brain. This was attributable to greater levels of both AT1A- and AT1B-R mRNA subtypes in these areas from the SHR. These observations suggest that increased AII receptor levels in SHR brain may, in part, be a result of increased expression of the AT1-R gene.     

Reduced periinfarction mortality as a result of long-term therapy with captopril but not hydralazine or propranolol in spontaneously hypertensive rats

Hypertension and left ventricular hypertrophy (LVH) are known to increase susceptibility to ventricular arrhythmias during and before myocardial ischemia and to increase the risk of periinfarction mortality. Although regression of LVH has been advocated as a therapeutic goal, little evidence exists to suggest that it can reduce periinfarction mortality, and if it does, by which mechanisms it may do this. In this study, we evaluated the effects of control of systemic arterial blood pressure, of regression of myocardial hypertrophy, and of cardiac fibrosis on the susceptibility to ventricular arrhythmias and periinfarction mortality in the spontaneously hypertensive rat (SHR) model of hypertension and LVH. After 12 weeks of treatment, captopril and hydralazine reduced systolic blood pressure to 93 +/- 14 and 126 +/- 13 mm Hg, respectively, as compared with 193 +/- 12 mm Hg, p < 0.05, in the untreated control SHR group. The decrease with propranolol (to 185 +/- 12 mm Hg) was of borderline significance. There was a significant decrease in inducibility of ventricular arrhythmias by programmed electrical stimulation with captopril (5%; p < 0.05). One hour after infarction, there was a trend toward reduced mortality in the rats treated with hydralazine, 9.5% (p = 0.20 vs. control; p = 0.10 vs. propranolol), and captopril, 5% (p = 0.08 vs. control; p = 0.010 vs. propranolol). However, only captopril reduced 3-h postinfarction mortality (40%; p = 0.022) compared with 72% in the control group. The results showed a significant decrease of the left ventricular weight/body weight ratio in the rats treated with hydralazine (2.6 +/- 0.2 mg/g; p < 0.05) and captopril (2.2 +/- 0.2 mg/g; p < 0.05) compared with the control group (2.8 +/- 0.2 mg/g). An assessment of cardiac fibrosis indicated that captopril decreased the volume percentage of collagen the most (2.01 +/- 0.53; p < 0.05), followed by propranolol (2.29 +/- 0.64; p < 0.05) and hydralazine (2.92 +/- 0.58; p < 0.05) versus controls (3.23 +/- 0.61). This study suggests that regression of myocardial hypertrophy or long-term normalization of arterial systolic blood pressure or both are the major determinants of very early mortality (within 1 h after infarction) and that later mortality (3 h after infarction) may be the result of a more complex interplay of regression of myocardial hypertrophy and fibrosis and of control of blood pressure.     

Evaluation of the alpha(2A)-adrenergic receptor gene in a heritable form of temporal lobe epilepsy

An autosomal dominant form of human temporal lobe epilepsy (TLE) has been mapped to a region of chromosome 10q that contains the intronless alpha(2A)-adrenergic receptor (alpha(2A)AR) gene. Because mutation of the alpha(2A)AR gene in the mouse fosters epileptogenesis, we developed methods for analysis of the alpha(2A)AR coding region applicable to any pathophysiologic state in which the alpha(2A)AR could be implicated in the disease mechanism. This study rules out mutations in the alpha(2A)AR coding region as causal for this form of autosomal dominant TLE.     

Dysfunctional D1A receptor-G-protein coupling in proximal tubules of spontaneously hypertensive rats is not due to abnormal G-proteins

Improving immunization coverage is vital to promoting child health and reducing childhood diseases and deaths. In spite of being actively promoted as a major public health intervention for national development since the late 1970s, immunization coverage in Ghana remains low. We investigated factors that influence attendance to immunization sessions in the Komenda-Edina-Eguafo-Abrem District of Ghana. The major factors hindering attendance were poor knowledge about immunization, lack of suitable venues and furniture at outreach clinics, financial difficulties, long waiting times, transport difficulties, poorly motivated service providers and weak intersectoral collaboration. The timing of immunization sessions, length of prior notice to the community, attitude of service providers and fear of side-effects generally did not deter attendance.