Richard H. Kennedy

Effects of streptozotocin-induced diabetes on heart rate, blood pressure and cardiac autonomic nervous control

Diabetes-associated alterations in resting heart rate and blood pressure have been demonstrated in clinical studies and in animal models of insulin-dependent diabetes mellitus (IDDM). These alterations may result from changes in the heart, vasculature or autonomic nervous system control. Using the streptozotocin- (STZ-) treated rat model of IDDM, the current study was designed to: (1) monitor changes in heart rate and blood pressure continually during a 10-week period in conscious unrestrained animals; and (2) determine if observed alterations in heart rate were mediated by changes in sympathetic and/or parasympathetic nervous control. Biotelemetry techniques were used. Heart rate and blood pressure were recorded 24 h a day at 10 min intervals before and after induction of diabetes. Diabetes was induced by i.v. administration of 50 mg/kg STZ. Resting autonomic nervous system tone was estimated by chronotropic responses to full-blocking doses of nadolol (5 mg/kg i.p.) and atropine (10 mg/kg i.p.). STZ-induced diabetes was associated with time-dependent reductions in heart rate and its circadian variation. Diastolic blood pressure and mean arterial pressure did not differ significantly when compared between control and STZ-treated animals; however, pulse pressure was diminished in diabetic rats. Chronotropic responses to both nadolol and atropine were blunted significantly in diabetic animals suggesting that resting levels of both vagal and sympathetic nervous tone to the heart were diminished. Heart rate in the presence of both nadolol and atropine was also decreased in diabetic rats. All effects observed following administration of STZ were reversed, at least in part, by insulin treatment. These results suggest that IDDM is associated with time-dependent reductions in resting heart rate and autonomic nervous control of cardiac function.

Burn Injury Alters the Intestinal Microbiome and Increases Gut Permeability and Bacterial Translocation

Sepsis remains one of the leading causes of death in burn patients who survive the initial insult of injury. Disruption of the intestinal epithelial barrier has been shown after burn injury; this can lead to the translocation of bacteria or their products (e.g., endotoxin) from the intestinal lumen to the circulation, thereby increasing the risk for sepsis in immunocompromised individuals. Since the maintenance of the epithelial barrier is largely dependent on the intestinal microbiota, we examined the diversity of the intestinal microbiome of severely burned patients and a controlled mouse model of burn injury. We show that burn injury induces a dramatic dysbiosis of the intestinal microbiome of both humans and mice and allows for similar overgrowths of Gram-negative aerobic bacteria. Furthermore, we show that the bacteria increasing in abundance have the potential to translocate to extra-intestinal sites. This study provides an insight into how the diversity of the intestinal microbiome changes after burn injury and some of the consequences these gut bacteria can have in the host.

Propofol-induced Alterations in Myocardial β-adrenoceptor Binding and Responsiveness

UNLABELLED Propofol (iv) depresses cardiovascular function in both humans and animals. However, the mechanism underlying this action has not been well described. The present study was designed to test the hypothesis that this effect of propofol results in part from an antagonism of adrenergic control of the heart. Experiments examined effects of propofol on: 1) [3H]CGP12177 (a beta-adrenoceptor antagonist) binding in rat myocardial membranes; and 2) the inotropic and chronotropic actions of isoproterenol in rat left atrial muscle and right atria, respectively. Propofol (25-200 microM) increased the apparent dissociation constant for [3H]CGP12177 without affecting binding site density. Similarly, 200 microM propofol increased the 50% effective concentration values for the dose-dependent positive chronotropic and inotropic actions of isoproterenol in right and left atria, and depressed the maximum increase in spontaneous rate elicited by this beta-adrenoceptor agonist. Other experiments demonstrated that propofol does not alter muscarinic receptor binding as monitored using [3H]quinuclidi-nylbenzilate. In conclusion, these results indicate that propofol can decrease cardiac beta-adrenoceptor responsiveness; however, the concentrations of propofol required suggest that this action contributes to the cardiovascular depression produced by this anesthetic only during large-dose bolus injection. IMPLICATIONS Experiments in membranes and cardiac preparations isolated from rat heart demonstrate that relatively high concentrations of propofol (25-200 microM) are required to antagonize beta-adrenoceptor binding and tissue responsiveness.

Inhibition of L-Type Ca2+ Channel Current in Rat Ventricular Myocytes by Terfenadine

To elucidate possible mechanisms underlying the cardiotoxicity of terfenadine, the effect of this antihistamine on L-type Ca2+ channel current (ICa,L) was studied in adult rat ventricular myocytes using the whole-cell patch-clamp technique. Myocytes were held at -70 mV and internally dialyzed and externally perfused with Na(+)- and K(+)-free solutions; exposure to terfenadine (10(-9) to 5 x 10(-6) mol/L) resulted in a concentration-dependent inhibition of peak ICa,L with a half-maximum inhibition concentration (IC50) of 142 nmol/L. The terfenadine-induced inhibition of ICa,L was not mediated via effects on histamine H1 receptors, because 1 mumol/L triprolidine, a more selective and potent H1 antagonist, had no effect on ICa,L. In this study, we found that terfenadine (1) increased both the fast and slow time constants of ICa,L inactivation, (2) shifted the steady state inactivation of ICa,L to more negative potentials, and (3) elicited a tonic block and a use-dependent block of ICa,L. The terfenadine-induced tonic and use-dependent block and the steady state inhibition of ICa,L were voltage dependent. Both tonic and use-dependent blocks of ICa,L by terfenadine at -40 mV were greater than that at -70 mV, and blocks were partially released by applying a long hyperpolarizing prepulse to -90 mV. These results suggest that terfenadine binds to L-type Ca2+ channels in inactivated and rested states and inhibits ICa,L predominantly by interacting with the inactivated state with an apparent dissociation constant of 60 nmol/L. Open-state block could be observed only at high concentrations of terfenadine. The high-affinity interaction of terfenadine with the inactivated state of L-type Ca2+ channels may play an important role in its cardiotoxicity under pathophysiological conditions, such as ischemia.

Influence of Mast Cells on Structural and Functional Manifestations of Radiation-Induced Heart Disease

Radiation-induced heart disease (RIHD), characterized by accelerated atherosclerosis and adverse tissue remodeling, is a serious sequelae after radiotherapy of thoracic and chest wall tumors. Adverse cardiac remodeling in RIHD and other cardiac disorders is frequently accompanied by mast cell hyperplasia, suggesting that mast cells may affect the development of cardiac fibrosis. This study used a mast cell-deficient rat model to define the role of mast cells in RIHD. Mast cell-deficient rats (Ws/Ws) and mast cell-competent littermate controls (+/+) were exposed to 18 Gy localized single-dose irradiation of the heart. Six months after irradiation, cardiac function was examined by echocardiography and Langendorff-perfused isolated heart preparation, whereas structural changes were assessed using quantitative histology and immunohistochemical analysis. Mast cell-deficient rats exhibited more severe postradiation changes than mast cell-competent littermates. Hence, mast cell-deficient rats exhibited a greater upward/leftward shift in the left ventricular (LV) diastolic pressure-volume relationship (P = 0.001), a greater reduction in in vivo LV diastolic area (from 0.50 +/- 0.024 cm in age-matched controls to 0.24 +/- 0.032 cm after irradiation; P = 0.006), and a greater increase in LV posterior wall thickness (from 0.13 +/- 0.003 cm in age-matched controls to 0.15 +/- 0.003 cm after irradiation; P = 0.04). Structural analysis revealed more pronounced postradiation accumulation of interstitial collagen III but less myocardial degeneration in hearts from mast cell-deficient rats. These data show that the absence of mast cells accelerates the development of functional changes in the irradiated heart, particularly diastolic dysfunction, and suggest that, in contrast to what has been the prevailing assumption, the role of mast cells in RIHD is predominantly protective.

Suppression of β-adrenergic responsiveness of L-type Ca2+ current by IL-1β in rat ventricular myocytes

The possible mechanism by which interleukin-1β (IL-1β) affects β-adrenergic responsiveness of L-type Ca2+ current ( I Ca,L) was examined in adult rat ventricular myocytes by use of whole cell patch-clamp techniques. In the presence of isoproterenol (Iso), exposure for 3 min to IL-1β suppressed the Iso-activated I Ca,L. In the presence of IL-1β, the response of I Ca,L to Iso was decreased, and the EC50 for Iso stimulation was increased. However, IL-1β had no effect on [3H]CGP-12177 binding, displacement of [3H]CGP-12177 binding by Iso, or on basal and Iso-enhanced cAMP content. When I Ca,L was activated by extracellular application of forskolin or 8-(4-chlorophenylthio)-cAMP, a membrane-permeable cAMP analog, or by intracellular dialysis with cAMP, IL-1β had little effect on I Ca,L. In contrast, in the presence of cAMP, IL-1β still suppressed the Iso-enhanced I Ca,L. These results show that the IL-1β-induced decrease in β-adrenergic responsiveness of I Ca,L does not result from inhibition of β-adrenoceptor binding, adenylyl cyclase activity, or cAMP-mediated pathways, suggesting a cAMP-independent mechanism.

Nitric oxide has no chronotropic effect in right atria isolated from rat heart

This study was designed to determine if nitric oxide (NO) has direct effects on heart rate or if it is involved in the chronotropic actions of adrenergic or cholinergic stimulation. Right atria were isolated from hearts of adult male rats, bathed in Krebs-Henseleit buffer (37 degrees C), and used to monitor spontaneous rate. For comparison, ring segments of thoracic aorta were also suspended in the Krebs-Henseleit solution and used to examine vascular actions of various agents. The dose-dependent chronotropic effects of acetylcholine (10(-7)-10(-3) M) and norepinephrine (10(-8)-3 x 10(-4) M) in right atria were not affected by pretreatment with 10(-4) M N-nitro-L-arginine or 10(-3) M N-nitro-L-arginine-methyl ester, inhibitors of L-arginine-derived NO production. SIN-1 (3-morpholino-sydnonimine), an agent which releases NO in aqueous solution, elicited a dose-dependent (0.3-100 microM) vasorelaxation in aortic preparations constricted with 60 mM KCl; the ED50 value for this effect was increased by pretreatment with methylene blue (10 microM) and LY-83,583 (6-(phenylamino)-5,8- quinolinedione; 1 and 3 microM), compounds which inhibit NO-induced stimulation of guanylate cyclase. SIN-1 produced a negative chronotropic effect in right atria; however, this action was not observed at concentrations less than 300 microM and was not antagonized by methylene blue or LY-83,583. 8-Bromo cyclic GMP produced a dose-dependent (10-3000 microM) decrease in KCl-induced tension in aortic rings. In right atria, 8-bromo cyclic GMP elicited a positive chronotropic effect.(ABSTRACT TRUNCATED AT 250 WORDS)

Gender differences in blood pressure and heart rate in spontaneously hypertensive and Wistar-Kyoto rats

1. In general, premenopausal women are known to have lower blood pressure than men and animal models have shown a similar sexual dimorphism. However, many studies in animals have been performed using anaesthetized or restrained models. Current experiments were conducted to investigate the relationships among resting heart rate, blood pressure and gender in conscious, unrestrained normotensive Wistar‐Kyoto (WKY) rats and spontaneously hypertensive rats (SHR).

Aging: changes in cardiac α1-adrenoceptor responsiveness and

Several investigators have reported a diminished responsiveness of senescent cardiac muscle to norepinephrine and beta-adrenoceptor agonists. In contrast, relatively little is known regarding the effects of aging on myocardial actions mediated specifically by alpha-adrenoceptor stimulation. Thus, the current study examined aging-dependent changes in: (a) the inotropic response to methoxamine, an alpha-adrenoceptor agonist; (b) characteristics of myocardial alpha 1-adrenoceptors as monitored by specific [3H]prazosin binding; and (c) steady state levels of alpha 1-adrenoceptor mRNA as determined by Northern blot analysis. Cardiac preparations were isolated from 4-, 14-, and 25-month-old F344 rats. An aging-associated decline was observed in the maximum positive inotropic effect elicited by methoxamine in right ventricular strips (160 +/- 23, 134 +/- 13 and 79 +/- 26% increase above control developed tension in 4, 14 and 25 months, respectively) with no change in ED50 values. [3H]Prazosin binding to ventricular sarcolemmal membranes revealed a reduction in receptor number (82 +/- 7, 69 +/- 6 and 59 +/- 5 fmol/mg protein in 4, 14 and 25 months, respectively); the apparent dissociation constant was not affected. Steady state levels of alpha 1-adrenoceptor mRNA decreased progressively between 4 and 25 months of age (14- and 25-month levels were approximately 71 and 38% of 4 months, respectively), while steady state levels of beta-actin mRNA did not change with age. These results suggest that the aging-related decline in alpha 1-adrenergic responsiveness in rat ventricular muscle is mediated, at least in part, by a decrease in cardiac alpha 1-adrenoceptor density.(ABSTRACT TRUNCATED AT 250 WORDS)

α1-Adrenergic activation of L-type Ca current in rat ventricular myocytes: perforated patch-clamp recordings

α1-Adrenergic stimulation has little effect on L-type Ca2+channel current ( I Ca,L) in adult cardiac myocytes measured using conventional whole cell voltage-clamp techniques. In this study using perforated-patch techniques, we reevaluated the effect of α1-adrenergic stimulation on I Ca,L in adult rat ventricular myocytes. Action potentials and I Ca,L were examined in the presence of 1 μM nadolol, a β-adrenergic antagonist, in myocytes internally dialyzed with Na+- and K+-free solutions (Cs+ and tetraethylammonium as substitutes). Phenylephrine (PE; 30 μM) increased the action potential duration measured at 25 and 70% of repolarization by 104 and 86%, respectively. In the perforated-patch configuration, PE elicited a transient decrease followed by a ∼60% increase in I Ca,L, whereas only the transient decrease in I Ca,L was observed in myocytes when the conventional whole cell configuration was used. The PE-induced increase in I Ca,L was reversibly blocked by 1 μM prazosin, an α1-adrenergic antagonist. These results suggest that α1-adrenergic stimulation enhances cardiac I Ca,L and that obligatory intracellular mediators for this action are lost during whole cell recordings.