A citywide evaluation of identification of risk factors for cardiovascular disease in emergency department patients complaining of chest pain.

The local division of the American Heart Association in Lubbock, Tex, conducted a cooperative study involving all three local hospitals to assess how effectively risk factors for cardiovascular disease are identified in patients presenting in the emergency departments with a complaint of chest pain. The charts of 250 consecutive patients with chest pain were reviewed for risk factors identified by the triage nurse and by the emergency department physician or the attending physician or both. Although the rate at which risk factors were identified was good, identification was neither complete nor comprehensive. Comparison with statistics for the general population showed that some risk factors were over-reported, while others were markedly underreported. Each of the three specialties of health care professionals stressed different risk factors, and having all three involved markedly increased the level of identification. The data provide guidelines for improving risk factor identification, and the study led to the formation of strategic alliances among the different hospitals and health care professionals that should help improve secondary prevention of cardiovascular disease within the community.

Benzylidene analogs of anabaseine display partial agonist and antagonist properties at the mouse 5-hydroxytryptamine(3A) receptor.

The nicotinic receptor drug candidate, 3-(2,4-dimethoxybenzylidene)-anabaseine (also known as GTS-21; DMXBA), its hydroxy metabolites, and some related analogs were evaluated with the two-electrode voltage-clamp technique in mouse 5-hydroxytryptamine (5-HT)(3A) receptors expressed in Xenopus oocytes. Although DMXBA lacked partial agonist activity, its hydroxy-benzylidene metabolites and related analogs were partial agonists, displaying the following rank order of potency (EC(50)) and apparent efficacy: 5-HT, 0.9 +/- 0.06 microM (100% efficacy) > 3-(2-hydroxy,4-methoxybenzylidene)-anabaseine (2-OH-MBA), 2.0 +/- 0.3 microM (63% efficacy) > 3-(2,4-dihydroxybenzylidene)-anabaseine, 2.6 +/- 0.3 microM (63% efficacy) > 3-(2-methoxy,4-hydroxybenzylidene)-anabaseine, 17.2 +/- 1.0 microM (30% efficacy). To examine the influence of a benzylidene ring hydroxy substituent, the agonist actions of the three possible monohydroxy isomers were examined. The rank order of potency, based on EC(50) determinations, and apparent efficacy was: 3-(2-hydroxybenzylidene)-anabaseine, 20.3 +/- 2.6 microM (63% efficacy) > 3-(4-hydroxybenzylidene)-anabaseine, 32.3 +/- 5.9 microM (14% efficacy) > 3-(3-hydroxybenzylidene)-anabaseine (3-OH-BA) (no agonist activity). Both DMXBA and 3-OH-BA antagonized 5-HT-mediated currents, with IC(50) values of 15.7 +/- 0.9 and 27.5 +/- 4.7 microM, respectively. DMXBA demonstrated both competitive and noncompetitive forms of antagonism over the range of concentrations tested. These results suggest that a hydroxy substituent at the 2' position of the benzene ring is necessary and sufficient for partial agonist activity; substitution at the 4' position with a hydroxy or methoxy group further enhances agonist potency. Because 2-OH-MBA is a primary metabolite of DMXBA, it may contribute to the physiological, biochemical, and behavioral effects of the parent compound when administered in vivo.

High-affinity interactions of ligands at recombinant guinea pig 5HT7 receptors.

The serotonin 5HT7 receptor has been implicated in numerous physiological and pathological processes from circadian rhythms to depression and schizophrenia. Clonal cell lines heterologously expressing recombinant receptors offer good models for understanding drug-receptor interactions and development of quantitative structure-activity relationships (QSAR). Comparative Molecular Field Analysis (CoMFA) is an important modern QSAR procedure that relates the steric and electrostatic fields of a set of aligned compounds to affinity. Here, we utilized CoMFA to predict affinity for a number of high-affinity ligands at the recombinant guinea pig 5HT7 receptor. Using R-lisuride as the template, a final CoMFA model was derived using procedures similar to those of our recent papers. The final cross-validated model accounted for >85% of the variance in the compound affinity data, while the final non-cross validated model accounted for >99% of the variance. Model evaluation was done using cross-validation methods with groups of 5 ligands. Twenty cross-validation runs yielded an average predictive r2(q2) of 0.779 +/- 0.015 (range: 0.669-0.867). Furthermore, 3D-chemical database search queries derived from the model yielded hit lists of promising agents with high structural similarity to the template. Together, these results suggest a possible basis for high-affinity drug action at 5HT7 receptors.

Competitive antagonism of the mouse 5-hydroxytryptamine3 receptor by bisindolylmaleimide I, a "selective" protein kinase C inhibitor.

We examined the effects of several protein kinase C (PKC) inhibitors on the murine 5-hydroxytryptamine3 (5-HT3) receptor to determine whether they acted directly on the receptor. The 5-HT-evoked currents in Xenopus laevis oocytes expressing the recombinant 5-HT3 receptor were measured with the two-electrode voltage-clamp technique. The PKC inhibitors bisindolylmaleimide I (BIM, GF109203x) and staurosporine, but not calphostin C or chelerythrine, decreased the 5-HT3 receptor-mediated currents when coapplied with 5-HT. BIM blocked 0.5 microM 5-HT-elicited currents with an IC50 value of 7 nM, whereas in the presence of 5 microM staurosporine, 42% inhibition of 0.5 microM 5-HT-mediated currents was observed. Increasing concentrations of BIM resulted in a rightward shift of the 5-HT concentration-response curve, without altering efficacy. A Schild plot was generated, which had a slope of -1.01, suggesting competitive antagonism. The Ki value of BIM was determined to be 29 nM. To confirm competitive antagonism, a competitive binding assay was performed on Sf21 insect cells infected with the mouse 5-HT3 receptor cDNA in a baculovirus expression vector. BIM completely displaced binding of the selective 5-HT3 receptor antagonist [3H]GR65630. BIM bound to the 5-HT3 receptor with a Ki value of 61 nM, which was slightly less potent than that of the selective 5-HT3 receptor antagonist MDL72222 (27 nM). The PKC inhibitor BIM is a potent competitive antagonist at the 5-HT3 receptor.

Inhibition of rat vascular smooth muscle cell proliferation by taurine and taurine analogues.

The growth of rat aorta vascular smooth muscle cells (VSMCs) was measured in the presence and absence of taurine. Concentrations of taurine as low as 0.3 mM in the culture medium inhibited the proliferation of the cells, as monitored by measuring cell count, and also inhibited the rate of DNA synthesis, as examined by measuring [3H]thymidine incorporation into DNA. However, even at the highest concentration of taurine (30 mM), the doubling time of the VSMCs was only increased by 38%. Protein content of the VSMCs was decreased by 30 mM taurine. [3H]Leucine incorporation into newly synthesized protein was not affected by the highest concentration of taurine tested (30 mM), indicating that taurine did not inhibit protein synthesis but rather decreased total protein content by inhibiting cellular proliferation. The effects of other amino acids such as alanine, glycine, and serine and of various taurine analogues such as beta-alanine, guanidinoethanesulfonic acid (GES), and isethionic acid also were tested at a concentration of 20 mM for their effects on the growth of the VSMCs. Alanine, glycine, and serine had only a minimal effect or no effect on cell count, quantity of protein, and incorporation of [3H]thymidine into DNA. GES, beta-alanine, and isethionic acid had a significant effect on cell count, protein content, and incorporation of [3H]thymidine into DNA. Beta-alanine was the only analogue tested that significantly depressed [3H]leucine incorporation into newly synthesized protein. It is concluded that taurine, GES, and isethionic acid inhibited proliferation of VSMCs but did not alter normal protein synthesis or survivability of VSMCs. In contrast, other amino acids, alanine, glycine and serine, had minimal effects on VSMC proliferation and protein synthesis, whereas beta-alanine appeared to be toxic, inhibiting both VSMC synthesis and de novo protein synthesis.

Alterations of rabbit aortic smooth muscle cell proliferation in diabetes mellitus.

OBJECTIVE: Diabetes mellitus is a known risk factor for atherosclerosis. Because initiation and/or progression of the atherosclerotic process is associated with alterations in vascular smooth muscle cell growth and differentiation, the present studies were conducted to evaluate the effect of diabetes mellitus on the proliferative behaviour of cultured aortic smooth muscle cells. METHODS: Male New Zealand White rabbits were made diabetic with a single intravenous injection of alloxan monohydrate (100 mg.kg-1) in saline. Primary cultures of smooth muscle cells were established from thoracic aortic segments of control and diabetic rabbits and used to develop multiple cell strains. The proliferative capability of secondary cultures was determined by measurements of [3H]-thymidine incorporation into DNA, cell counts, and protein content in control and diabetic cultures. The serum dependence of cellular growth was evaluated by incubation of cultured cells in growth medium supplemented with various fetal calf serum concentrations. RESULTS: Cultures of diabetic origin incorporated thymidine to a greater extent than control cultures. Although the efficiency of cell attachment was not different between control and diabetic cells, diabetic cells had a shorter population doubling time than control cells [41.08(SEM 4.15) h v 58.08(6.79) h] and achieved higher final densities than control cultures. The serum dependence of smooth muscle cell cultures for viability and growth was different between the two groups. CONCLUSIONS: These findings support the hypothesis that diabetes induces changes in vascular smooth muscle cell proliferation which may be associated with the onset or progression of the atherogenic process observed in diabetes.

Changes in cytoplasmic pH in platelets activated through the high- and moderate-affinity receptor pathways by highly purified human alpha-thrombin.

Platelets in plasma were loaded with the probe BCECF/AM, and changes in cytoplasmic pH levels induced by highly purified human alpha-thrombin (2900 NIH U/mg) were studied in washed platelets having high- and moderate-affinity receptors and in platelets from which the high-affinity alpha-thrombin receptor had been removed by treatment with Serratia marcescens protease. In intact platelets, cytoplasmic acidification reached a maximum within 2 minutes of -0.072 +/- 0.009 pH units at 0.3 nmol/L alpha-thrombin concentration (0.03 U/ml). Cytoplasmic pH values were higher at both lower and higher alpha-thrombin concentrations and were significantly (p = 0.018) higher at 2 nmol/L alpha-thrombin, which induced -0.037 +/- 0.013 pH units of acidification. Five nanomoles of alpha-thrombin, however, induced cytoplasmic alkalinization of +0.027 +/- 0.033 pH units. In platelets lacking the high-affinity receptor where there is a 10 to 20-fold reduction in sensitivity to alpha-thrombin, acidification reached a maximum of -0.175 +/- 0.033 pH units at 2 nmol/L alpha-thrombin, but alkalinization was observed at 5 nmol/L (+0.038 +/- 0.025) and 10 nmol/L (+0.042 +/- 0.007) alpha-thrombin. These results show that the transition from acidification to alkalinization occurs in the same range of alpha-thrombin concentrations (2 to 5 nmol/L) in both preparations, despite the rightward shift in sensitivity caused by the absence of the high-affinity receptor. However, the maximum acidification reached in control platelets (-0.037 pH units at 2 nmol/L) was much less than the value obtained in platelets lacking the high-affinity receptor (-0.175 pH units at 2 nmol/L alpha-thrombin).(ABSTRACT TRUNCATED AT 250 WORDS)

Pharmacological study of isoproterenol and diabetic cardiomyopathies in rat right ventricular strips.

The present study was undertaken to compare the cardiomyopathies induced by experimental diabetes mellitus and chronic isoproterenol (ISO) pretreatment on inotropic reactivity of right ventricular strips from rats. One month after initiation of treatment, cardiac muscle was isolated and challenged with either the beta-adrenoceptor agonist ISO or the calcium channel activator Bay K 8644. Sensitivity and responsiveness to these agonists were determined. Crude membrane preparations were obtained from residual ventricular muscle and beta-adrenoceptor density determined using iodocyanopindolol. Isolated right ventricular strips from diabetic and ISO-treated rats demonstrated hyperresponsiveness to the inotropic effects of Bay K 8644 but subsensitivity to ISO. Ventricular beta-adrenoceptor density was found to be significantly depressed in membrane preparations from both diabetic and ISO-treated groups relative to controls. These data indicate that both forms of cardiomyopathy may affect inotropic reactivity through similar alterations to beta-adrenoceptors and calcium utilization.

Phencyclidine binds to blood platelets with high affinity and specifically inhibits their activation by adrenaline.

The ion channel probe phencyclidine [1-(1-phenylcyclohexyl)piperidine; PCP] selectively inhibited aggregation, secretion and ultrastructural changes in platelets induced by adrenaline, but did not affect activation induced by other common platelet agonists such as alpha-thrombin, ADP, collagen or ionophore A23187. [3H]PCP bound to platelets with high affinity (Kd 134 +/- 33 nM; 3600 +/- 1020 sites/platelet), as did the thienyl analogue [3H]TCP (1-[1-(2-thienyl)cyclohexyl]piperidine). PCP binding to platelets was increased 3-4-fold in N-methylglucamine buffer in the absence of Na+ ions. Binding was unaffected by haloperidol and was only weakly inhibited (EC50 10-20 microM), without significant stereoselectivity by the two sets of stereoselective ligands, dexoxadrol/levoxadrol and (+)MK801/(-)MK801. Binding of PCP was not competed for by adrenaline or yohimbine. Only the high-affinity binding of [3H]PCP to platelets was blocked by prior treatment of the platelets with the covalent affinity probe Metaphit, and these platelets no longer aggregated in response to adrenaline although they responded normally to alpha-thrombin, ADP and collagen. These results suggest that platelets contain high-affinity receptors for PCP that can modulate adrenaline-induced platelet activation.

Rabbit aortic smooth muscle cell culture. A model for the pharmacological study of diabetes-induced alterations in cell proliferation.

Atherosclerotic vascular disease is the most common complication of diabetes mellitus. Enhanced vascular smooth muscle cell proliferation plays a central role in atherosclerotic lesion formation. Studies using explant cultures have demonstrated that aortic smooth muscle cells from rats with experimental or genetic diabetes have enhanced rates of proliferation when compared to controls. However, this method of culture may select for cells with enhanced migratory potential. In the present studies, aortic smooth muscle cells were successfully cultured from control and diabetic rabbits after enzymatic and mechanical dispersion from thoracic aortic segments. The proliferative patterns of control cells were characterized and growth rates of diabetic cells were compared to controls. Primary cultures from control rabbits grew after an initial 5-day lag period to achieve threefold increases in cell number by 9 days. Subcultures of aortic smooth muscle cells entered the logarithmic phase of growth after 2 days, reaching the plateau phase of growth in 5-7 days and achieving three to fourfold increases in cell number. The final density to which cultures grew was not affected by the number of cells attached on day 1 for the range studied. Cells from diabetic rabbits displayed shorter doubling times and reached greater densities at confluence than did cells from controls. These data support the hypothesis that diabetes induces an atherogenic response. The dissociated rabbit aortic smooth muscle cell culture provides a model in which to study diabetes-induced modulation of cell proliferation that is amenable to pharmacological manipulation to investigate agonist and growth factor-induced responses.

PPACK-thrombin inhibits thrombin-induced platelet aggregation and cytoplasmic acidification but does not inhibit platelet shape change.

We have re-evaluated the previously reported ability of TLCK-thrombin (N alpha-tosyl-L-lysine chloromethyl ketone-treated alpha-thrombin) and PPACK-thrombin (D-phenylalanyl-L-prolyl-L-arginine chloromethyl ketone-treated alpha-thrombin) to inhibit alpha-thrombin-induced platelet activation (Harmon JT, Jamieson GA: J Biol Chem 261:15928, 1986; and Harmon JT, Jamieson GA: Biochemistry 27:2151, 1988). Despite several cycles of derivatization with TLCK (10,000-fold molar excess), preparations of TLCK-thrombin have been found to contain about 4% residual alpha-thrombin activity, suggesting that these preparations are an equilibrium mixture of TLCK-thrombin and alpha-thrombin and cannot be used for evaluating competition between these two agents. In contrast, alpha-thrombin activity was completely inhibited by PPACK at 15-fold molar excess. PPACK-thrombin, free of unreacted PPACK and devoid of residual alpha-thrombin activity, did not markedly affect platelet shape change at concentrations as high as 1 mumol/L, but inhibited aggregation and secretion in intact platelets activated with the minimal concentration of alpha-thrombin causing a full response (0.3 to 0.5 nmol/L) and yielded a 50% inhibition constant (IC50) for inhibition of aggregation by PPACK-thrombin of 110 nmol/L. This inhibition was specific for alpha-thrombin-induced platelet activation, and no inhibition was seen with activation induced by ADP, collagen, epinephrine, ristocetin, or arachidonate. At these low alpha-thrombin concentrations (approximately 0.4 nmol/L), a persistent cytoplasmic acidification was observed of -0.062 +/- 0.016 pH units, although alkalinization was observed at higher alpha-thrombin concentrations (greater than 1 nmol/L). While inhibition of aggregation and secretion occurred when alpha-thrombin and PPACK-thrombin were added simultaneously, inhibition of cytoplasmic acidification and of the elevation of cytoplasmic [Ca2+] induced by low concentrations of alpha-thrombin (0.4 nmol/L) occurred only if platelets were preincubated with PPACK-thrombin for 5 minutes before the addition of alpha-thrombin. In platelets treated with Serratia marcescens protease to remove glycoprotein lb (GPlb), alpha-thrombin-induced shape change was attenuated but persisted in the presence of a high concentration (2 mumol/L) of PPACK-thrombin, although aggregation and secretion were inhibited, as seen in intact platelets. The IC50 value for inhibition of aggregation by PPACK-thrombin was approximately 1 mumol/L at the higher alpha-thrombin concentrations (5 nmol/L) required for full activation in this case. These results suggest that PPACK-thrombin may be a useful probe of platelet function since it specifically blocks platelet aggregation and secretion induced by alpha-thrombin.(ABSTRACT TRUNCATED AT 400 WORDS)

Functional characterization of beta-adrenoceptor subtypes in rabbit right atria.

The advent of radioligand binding studies has allowed the classification of receptor subtypes in various tissues. However, the presence of a receptor subtype in a heterogenous tissue does not insure that the receptor has a significant physiological role. beta 1- and beta 2-Adrenoceptors have been reported to coexist in the rabbit right atria. The purpose of the present investigation was to determine the physiological role of beta-adrenoceptor subtypes in catecholamine-induced chronotropic responses in the rabbit right atria through comparison of data from functional and radioligand binding studies. Rank order of potency was determined using isoproterenol, epinephrine and norepinephrine for both chronotropic and inotropic responses in the rabbit right atria and right ventricular papillary muscles, respectively. These studies indicated that the beta 1-adrenoceptor was primarily responsible for catecholamine-induced responses. Next, the beta 1-selective antagonist, atenolol, was found to inhibit the chronotropic responses of the nonselective beta-agonist, isoproterenol, and the beta 2-selective agonist, terbutaline, to the same extent. These data indicate that terbutaline produces its chronotropic effects in the rabbit right atria through stimulation of beta 1-, not beta 2-adrenoceptors. Finally, competition studies for [125I]iodocyanopindolol and the relatively selective beta 1- and beta 2-adrenoceptor antagonists (ICI 89406 and ICI 118551, respectively) indicated that the ratio of beta 1- to beta 2-adrenoceptor subtypes is 6:1. It is concluded that while both receptors may be present in the rabbit right atria, the beta 1-adrenoceptor is the predominant subtype both in density and physiological significance, while the beta 2-adrenoceptor plays little, if any role, in the chronotropic responses induced by catecholamines.

Characterization of beta-adrenoceptor subtypes in rabbit mononuclear leukocytes.

Computer analysis of [125I]iodocyanopindolol competition studies using the relatively selective beta 1-adrenoceptor antagonist, ICI 89406, and the beta 2-selective antagonist, ICI 118551, on rabbit mononuclear leukocyte plasmalemmal preparations favored a two-site model indicating that both beta 1- and beta 2-adrenoceptor subtypes were present in approximately equal numbers. In contrast, similar studies performed on rabbit cardiac sarcolemma favored a one site model consistent with the presence of beta 1-adrenoceptors. Consequently, rabbit mononuclear leukocytes may provide a useful model for studying selective modulatory mechanisms of beta 1- and beta 2-adrenoceptors.

Dietary calcium and development of hypertension in spontaneously hypertensive rats.

The purpose of the present study was to attempt to correlate four calcium diets (0.02, 0.1, 0.5 and 2.5%) with changes in the development of hypertension in both spontaneously hypertensive and Wistar-Kyoto rats. Our findings confirm that an inverse relationship exists between dietary calcium content and the development of hypertension. The relationship does not rely upon altered serum ionized sodium, potassium, or calcium or parathyroid hormone levels. In addition, no consistent dietary calcium-dependent changes were noticed in cardiovascular reactivity. In contrast, anesthesia with pentobarbital completely abolished the relationship. These data support the hypothesis that dietary calcium influences autonomic tone through some, as yet, undefined processes.

Nonspecific supersensitivity induced by reserpine in guinea pig cardiac ventricle tissue.

The depletion of norepinephrine stores by chronic reserpine pretreatment is thought to be responsible for the development of supersensitivity in cardiac tissue. The present study was designed to determine if reserpine-induced supersensitivity could be demonstrated in isolated guinea pig right ventricular strips, if it was associated with beta adrenoceptor proliferation and was specific for beta adrenoceptor agonists. In addition, reserpine dose-dependency of the phenomenon was tested for by using two pretreatment regimens to determine if supersensitivity was the result of direct or possibly toxic effects of reserpine. Pretreatment of guinea pigs with reserpine (0.1 mg/kg/day) for 7 days resulted in over 90% depletion of cardiac norepinephrine stores. Supersensitivity to the inotropic effects of isoproterenol, impromidine and forskolin was demonstrated. Associated with the supersensitivity was a significant increase in beta adrenoceptor density. Muscarinic receptor density was actually decreased. This pretreatment regimen also was associated with a significant (30%) drop in body weight suggesting that the phenomenon might be the result of a direct toxic effect of reserpine. Pretreatment of guinea pigs with a lower dose of reserpine (0.03 mg/kg/day) for 7 days produced the same degree of norepinephrine depletion, nonspecific inotropic supersensitivity and beta adrenoceptor proliferation in the absence of a significant reduction in body weight. The degree of supersensitivity induced by the two pretreatment regimens was between 2- to 3-fold for all three agonists tested.(ABSTRACT TRUNCATED AT 250 WORDS)

Electromechanical effects of bPTH-(1-34) on rabbit sinus node cells and guinea pig papillary muscles.

The effects of bPTH-(1-34), a synthetic preparation of bovine parathyroid hormone containing the first 34 amino acids, on electromechanical activity of isolated rabbit sinus node cells and guinea pig papillary muscles were examined by microelectrode techniques. In sinus node cells, bPTH-(1-34) (10(-7) M) decreased the cycle length of spontaneous firing (SPCL) accompanied by an increase in the maximum upstroke velocity at phase 0 (dV/dtmax) without affecting the amplitude of the action potential (APA) or the maximum diastolic potential (MDP). All the effects of bPTH-(1-34) on sinus node cells were abolished by verapamil (5 X 10(-7) M), but not by pindolol (2 X 10(-7) M). In constantly driven guinea pig papillary muscles, bPTH-(1-34) caused a positive inotropic effect (+31%). The parameters of the action potential were not significantly affected. This inotropic effect of bPTH-(1-34) was inhibited by pretreatment with verapamil or a low calcium medium (0.12 mM), but was not affected by pindolol. In contrast, ryanodine (2 X 10(-6) M), an inhibitor of internal Ca2+ release, which decreased the contraction with a prolongation of the action potential duration augmented the inotropic effect of bPTH-(1-34). In papillary muscles depolarized by 26 mM [K+]O, bPTH-(1-34) enhanced the slow action potential. In voltage clamp experiments using a single sucrose-gap method, bPTH-(1-34) caused an increase in the peak amplitude of the slow inward current, while it did not affect the outward current.(ABSTRACT TRUNCATED AT 250 WORDS)

Determination of beta adrenoceptor density on rabbit mononuclear leukocytes.

The purpose of the present study was to devise a technique for the isolation of a relatively homogeneous mononuclear leukocyte (MNL) preparation from rabbit whole blood and determine the density and affinity of beta-adrenoceptors on MNL. A modified method based upon that by Böyum was developed to maximize isolation of MNL from red blood cells (RBC), granulocytes, and platelets. The method involved an initial centrifugation to remove platelets and two centrifugations with a Ficoll solution to eliminate RBC and granulocytes. beta-Adrenoceptor density as determined with [125I]cyanopindolol and MNL membrane or whole cell preparations ranged between 317 and 360 sites per cell. Affinity for the binding sites was dependent upon whether membrane or whole cell preparations were studied, being 56.3 +/- 9.9 and 11.4 +/- 1.4 pM, respectively. Binding sites were found to be saturable and noncooperative. In addition, the binding sites demonstrated selectivity and stereospecificity for beta-adrenoceptor ligands. It is concluded that the modified method of harvesting MNL from rabbit whole blood provides a relatively homogeneous cell suspension that can be used to study the beta-adrenoceptor system.

Alterations in the myocardial beta-adrenoceptor system of streptozotocin-diabetic rats.

Previous investigations in our laboratory revealed subsensitivity of right ventricular tissue, isolated from one month STZ-diabetic rats, to the inotropic effects of isoproterenol. The present study was concerned with the characterization of this subsensitivity phenomenon. Observations of supersensitivity to methoxamine accompanied by decreased responsiveness to glucagon without a change in responsiveness to forskolin suggested a specific effect of diabetes on pathways involving receptor-mediated activation of adenylate cyclase. Radioligand binding analysis further revealed a specific decrease in the population of the high affinity state of the beta-adrenoceptor. Since the high affinity receptor state is a necessary intermediate for adenylate cyclase activation and enhanced myocardial contractility, it is proposed that the specific decrease in the high affinity population of the beta-adrenoceptor contributes to myocardial subsensitivity to isoproterenol observed in the diabetic animals. It is further proposed that the decrease in receptor population is related to increases in circulating epinephrine levels which were evident in the diabetic animals.

Prevention of streptozotocin-induced alterations in the rat heart by 3-O-methyl glucose and insulin treatments.

Streptozotocin-induced diabetes has previously been shown to alter the sensitivity and responsiveness of rat myocardial tissues to cardiotonic agonists. The objective of the present study was to determine if these alterations were due to the diabetogenic or possible direct cardiotoxic effects of streptozotocin. One month after streptozotocin treatment the following changes were observed in the rat: decrease in body weight; elevation of blood glucose and glycosylated hemoglobin levels; decrease in spontaneously beating atrial rate; elevation in basal developed force of electrically driven right ventricle; and inotropic subsensitivity of right ventricle to isoproterenol, which was associated with decreased beta-adrenoceptor density and supersensitivity to calcium. Pretreatment with the nonmetabolizable glucose analog 3-O-methyl glucose prevented these alterations. Chronic insulin replenishment also reversed the effects of streptozotocin, with the exception of complete normalization of elevations in blood glucose and basal developed force. Acute exposure to high glucose in the medium preserved the subsensitivity to isoproterenol but resulted in an elevated basal developed force in both control and streptozotocin groups. These observations indicate that myocardial alterations after streptozotocin treatment are not the result of direct cardiotoxic effects but rather a consequence of the drug-induced diabetic state. They also suggest that the increase in basal developed force might be related to elevated glucose concentrations.