Regulation of inositol 1,4,5-trisphosphate metabolism by guanine nucleotides in membranes of cultured newborn rat cardiomyocytes.

Membranes of cultured newborn rat cardiomyocytes contain enzymatic activities that regulate the formation and the breakdown of inositol 1,4,5-trisphosphate (1,4,5-IP3). GTP gamma S increased the rate of exogenous [3H]phosphatidyl 4,5-bisphosphate ([3H]PIP2) hydrolysis (EC50: 40 microM). This effect was dependent on the presence of deoxycholate and maximal at 2 mM deoxycholate. GTP gamma S increased the efficacy of phospholipase C (PLC) (by 2.3-fold), but did not alter the apparent affinity of the enzyme for PIP2. Other nucleotides, GDP beta S and ATP gamma S, and pyrophosphate also stimulated PIP2 hydrolysis, while AlF4- was ineffective. The effect of GTP gamma S was not inhibited by GDP beta S. The agonists norepinephrine and thrombin, which by themselves had no effect, did not potentiate the response to GTP gamma S. In contrast, 1,4,5-IP3 hydrolysis was decreased by GTP gamma S (EC50: 100 microM) as well as by other nucleotides and by pyrophosphate, but not by AlF4-. GDP beta S did not antagonize the GTP gamma S-induced inhibition of IP3 hydrolysis. These results suggest that GTP can stimulate the hydrolysis of exogenous PIP2 by an action on membrane-bound PLC at a site beyond the G protein activating PLC and inhibit the hydrolysis of 1,4,5-IP3 by a mechanism common to all nucleotides. Thus, GTP can regulate 1,4,5-IP3 metabolism by stimulating its formation and inhibiting its breakdown.

Plasma neuropeptide Y and catecholamines in pediatric patients undergoing cardiac operations.

OBJECTIVE: Our objective was to assess the sympathoadrenal response in pediatric patients undergoing repair of congenital cardiac defects. METHODS: Plasma catecholamine (norepinephrine and epinephrine) and neuropeptide Y concentrations were quantified before and after cardiopulmonary bypass to assess the response to cardiopulmonary bypass. To determine the response to aortic occlusion, levels of plasma catecholamines and neuropeptide Y were measured at the time of and immediately after release of the aortic crossclamp. RESULTS: During cardiopulmonary bypass, no significant change in levels of plasma norepinephrine (n = 43), epinephrine (n = 37), or neuropeptide Y (n = 46) was observed. Aortic occlusion induced a significant increase in plasma neuropeptide Y, but not in catecholamines. There was a greater increase in plasma neuropeptide Y in children older than age 1 year than in those younger than 1 year. CONCLUSIONS: Plasma neuropeptide Y may be a useful marker of sympathetic nervous system activity. Children younger than age 1 year showed a lesser sympathetic response compared with the response in older children.

Interaction of the alpha-2 adrenergic- and opioid receptor with the cGMP system in the mouse cerebellum.

The alpha-2 adrenergic agonist dexmedetomidine (Dex), 3-300 microg/kg, i.p., decreased cerebellar cGMP in a dose-dependent manner. Fentanyl (F), an opioid agonist, increased cerebellar cGMP at 0.3 mg/kg, s.c., and decreased it at doses >/=1 mg/kg. The inhibitory effect was receptor specific, that of Dex being blocked by the alpha-2 adrenergic antagonist yohimbine, 5 mg/kg, i.p.; that of F by the opioid antagonist naloxone, 5 mg/kg, i.p. In contrast the stimulatory effect of F was blocked by both naloxone and yohimbine. Yohimbine also enhanced the inhibitory effect of F. In mice pretreated with pertussis toxin, 2 microgram/mouse, given i.c.v. 72 h before the agonists, the decrease in cGMP induced by Dex or F was not affected, while the stimulatory effect of F was reversed to an inhibitory effect. When inhibiting doses of F and Dex were administered together, the cGMP response was smaller than the sum of the individual responses. Dex attenuated in a dose-dependent manner the decrease in cGMP induced by F, and unmasked or enhanced the stimulatory effect of F. These results show that the alpha-2 adrenergic- and opioid-receptors are coupled to the cGMP effector system and suggest that the two pathways converge at a common post-receptor site in the cascade of events transducing the receptor signal to cGMP regulation.

The nitric oxide-cGMP system of the locus coeruleus and the hypnotic action of alpha-2 adrenergic agonists.

Dexmedetomidine (Dex), a potent, selective alpha-2 adrenergic agonist, injected bilaterally directly into the locus coeruleus (LC), 7 microgram/LC, or ip, 100 microgram/kg, produced a maximum decrease in LC cGMP (-38 and -46%, respectively). Atipamezole, a selective alpha-2 adrenergic antagonist, given into the LC, 10 microgram/LC, prevented the decrease in LC cGMP induced by Dex, given i.p. or into the LC. Dex produced a loss of righting reflex in about 80% of the animals, an effect which was prevented by pretreatment with atipamezole. The nitric oxide synthase antagonist L-Name injected bilaterally into the LC, 20 microgram/LC, produced by itself a maximum decrease in LC cGMP of 54.5%. Dex, 100 microgram/kg, ip, given after L-Name, did not further decrease LC cGMP. L-Name, by itself, did not produce a loss of righting reflex, while 6 out of 9 rats pretreated with L-Name lost their righting reflex in response to Dex. A role of the nitric oxide-cGMP system of the LC in the modulation of the hypnotic effect of alpha-2 adrenergic agonists remains uncertain.

Dexmedetomidine decreases extracellular dopamine concentrations in the rat nucleus accumbens.

The effects of dexmedetomidine, a highly selective alpha(2)-adrenoceptor agonist, on extracellular dopamine (DA) concentrations in the nucleus accumbens of awake rats were collected via in vivo cerebral microdialysis and measured using HPLC with electrochemical detection. The administration of dexmedetomidine (DEX) at a low dose (2 microg/kg bolus i.v. over 2 min followed by a continuous infusion of 0.1 microg/kg per min) and a high dose (20 microg/kg bolus i.v. over 2 min followed by a continuous infusion of 1 microg/kg per min), significantly decreased extracellular dopamine concentrations in the nucleus accumbens. The observed decrease was dose-dependent, occurring sooner and to a greater magnitude in the rats receiving a high dose of DEX. This inhibitory modulation of accumbal dopamine was receptor-specific, as the decrease in extracellular DA produced by DEX was no longer evident following pre-treatment and co-infusion with the highly selective alpha(2)-adrenoceptor antagonist, atipamezole (ATZ). Thus, these data suggest that adrenoceptor agonists and antagonists may modulate dopaminergic neurotransmission via mechanisms that are specific to the alpha(2)-adrenoceptor.

The myocardial depressant effect of volatile anesthetics does not involve arachidonic acid metabolites or pertussis toxin-sensitive G proteins.

The involvement of the phospholipid-arachidonic acid pathway and of a pertussis toxin-sensitive G protein in the myocardial depressant effect of volatile anesthetics was examined in the rat heart left papillary muscle and atria. Neither phospholipase A2, cyclooxygenase or lipoxygenase antagonists altered the potency or the efficacy of clinically used concentrations of halothane and isoflurane in decreasing the contractile force of the muscles. Pretreatment of rats with pertussis toxin (50 micrograms/kg i.v., 72 h before sacrifice), which abolished the myocardial depressant effect of muscarinic agonists, did not prevent the decrease in contractile force induced by the anesthetics. The results of this study indicate that the cellular mechanism of the myocardial depressant effect of volatile anesthetics does not involve a metabolite of the phospholipid-arachidonic acid pathway, does not require a functional pertussis toxin-sensitive G protein, and differs from that of muscarinic agonists.

Cyclic 3',5'-adenosine monophosphate and bronchial tone.

The present studies demonstrate that adenylate cyclase and cyclic 3',5'-adenosine monophosphate (cAMP)-phosphodiesterase activities in dog bronchus are comparable to those found in other smooth muscle preparations. Catecholamines, in the order isoproterenol greater than epinephrine greater than norepinephrine, increase the rate of cAMP formation. This effect can be competitively inhibited by propranolol and potentiated by a cAMP-phosphodiesterase inhibitor. The kinetic study of bronchial cAMP-phosphodiesterase showed two different rates of cAMP hydrolysis, with apparent Km values of 1.4 and 48.0 muM. The high affinity cAMP-phosphodiesterase was inhibited competitively by theophylline and papaverine, the latter being about 20 times more potent than the former. The potency of each compound to inhibit the enzyme and to relax the bronchial strip was comparable. These results, the similar order of potency of the catecholamines to relax the bronchus and to increase the rate of cAMP formation, the competitive inhibition of both effects by propranolol, and the relaxing effect of dibutyryl cAMP on bronchial strip, are compatible with the assumption that the cAMP system is one of the biochemical mechanisms mediating bronchial smooth muscle relaxation.

Halothane effect on cAMP generation and hydrolysis in rat brain.

The volatile anesthetic halothane increased the rate of cAMP generation and decreased the rate of cAMP hydrolysis in rat cerebral cortex and cerebellum. The effect of halothane on the enzymes was reflected in a two-fold rise in cAMP content of cerebral cortical tissue exposed to the anesthetic at 3 vol% for 15 and 30 min. The action of halothane on adenylate cyclase is calcium-independent and different from the action of guanine nucleotides, sodium fluoride and specific transmitters. The Vmax of the enzyme is higher in the presence of the anesthetic. It is suggested that halothane, through conformational changes of the enzyme, renders more catalytic sites operative.

Superselective intraarterial papaverine administration: effect on regional cerebral blood flow in patients with arteriovenous malformations.

In this study the authors determined the effect of papaverine on regional cerebral blood flow (rCBF) in the angiographically normal arteriolar beds of patients with arteriovenous malformations (AVMs) who underwent transfemoral superselective angiography. Middle cerebral artery (MCA) branch vessels were catheterized during 10 procedures performed in nine patients. The mean (+/- standard deviation) largest AVM diameter was 4.4 +/- 1 cm. Regional CBF was measured by recording the washout of a bolus of xenon-133 injected through the microcatheter. In a dose-ranging study. rCBF and MCA pressure in two patients were repeatedly measured after 3-minute infusions of papaverine at 0.07, 0.7, and 7 mg/minute. In a single-dose study, an additional eight patients received only the highest dose of papaverine administered over a 3-minute period. In the dose-ranging study, CBF increased from baseline in a dose-dependent fashion. In the single-dose study, papaverine increased in rCBF 103%, from 48 +/- 11 to 95 +/- 23 ml/100 g/minute at an MCA pressure of 55 +/- 23 mm Hg. Increase in rCBF was linearly related (y = 2.2x - 17, r2 = 0.84; p = 0.001) to baseline MCA pressure (range 22-84 mm Hg). Papaverine increases rCBF in a direct proportion to baseline MCA pressure, even at low baseline pressures. Selective infusion of vasodilators should be investigated in acute cerebral hypotension to facilitate either primary or collateral recruitment of CBF by aiding spontaneous autoregulatory vasodilation. In addition, rCBF monitoring may be useful in determining the most effective intraarterial dose of papaverine while minimizing complications due to hyperemia.

The nitric oxide-cyclic 3',5'-guanosine monophosphate signal transduction pathway in the mechanism of action of general anesthetics.

(1) The nitric oxide-cyclic 3',5'-guanosine monophosphate (NO-cGMP) system is a major signaling transduction pathway implicated in a wide range of physiologic and pathophysiologic functions of the cardiovascular, respiratory, gastrointestinal, nervous or immune systems. (2) Evidence is provided that, at anesthetic concentrations, volatile and intravenous anesthetics interact with the NO-cGMP system. They have been shown to produce a decrease in cGMP in neuronal and vascular tissue. (3) Inhibition of NO synthesis produces a dose-dependent reversible decrease in the minimum anesthetic requirement and in the ED50 for the loss of righting reflex induced by general anesthetics. Volatile anesthetics also inhibit the NO-mediated relaxation in many vascular beds. (4) The selective alpha-2 adrenergic agonist, dexmedetomidine, which has potent sedative/hypnotic, anesthetic sparing and analgesic properties, produces a dose-dependent, reversible decrease in cGMP in mouse cerebellum at concentrations that decrease the anesthetic requirement of volatile anesthetics or induce a loss of righting reflex, an effect eliminated when NO synthase is inhibited. The site and mechanism by which the anesthetics interact with the NO-cGMP system is not yet clear and may vary with the anesthetic.

Halothane interaction with guanine nucleotide binding proteins in mouse heart.

Volatile anesthetics have been shown to decrease hormone-induced adenosine cyclic monophosphate (cAMP) formation and to increase guanosine cyclic monophosphate (cGMP) content in mouse ventricular myocardium. Hormone-induced inhibition of adenylate cyclase, the enzyme that synthesizes cAMP, and the cGMP response to alpha adrenergic agonists are mediated by a guanine nucleotide binding protein (N) sensitive to pertussis toxin. To evaluate the involvement of N proteins in the action of halothane on cyclic nucleotides in the heart, mice were pretreated with pertussis toxin, 50 micrograms/kg, ip, 72 h prior to exposure to halothane, 1.2 vol%. Pretreatment with the toxin decreased the cGMP response to halothane by 65% but was without effect on the decrease in myocardial cAMP induced by the anesthetic. The results indicate that a functionally active pertussis toxin-sensitive N protein is involved in the cGMP response to halothane, but not in the cAMP response.

Effect of clonidine on myocardial cyclic GMP content in the mouse-activation of central and peripheral alpha adrenoceptors.

Clonidine (0.23-3.77 mumol/kg i.p.) produced a dose-dependent increase in mouse myocardial cyclic GMP (cGMP) content. This effect was antagonized by yohimbine (0.03-1 mg/kg i.p.), but not by prazosin (1 mg/kg i.p.). The inhibition by yohimbine was biphasic. The cGMP response to clonidine was inhibited by atropine (5 mg/kg i.p.) and methylatropine (0.2-5 mg/kg i.p.). In mice pretreated with the ganglionic blocker hexamethonium, the cGMP response to clonidine persisted. St-91 [(2,6-diethylphenylamino)-2-imidazoline] (0.39-3.94 mumol/kg i.p.), a cogener of clonidine which does not cross the blood-brain barrier, also increased myocardial cGMP content. The potency of clonidine was similar in mice pretreated and nonpretreated with hexamethonium. Methylatropine did not affect the cGMP response to St-91 and to clonidine in ganglionectomized mice and yohimbine was a less potent antagonist. These results indicate that systemic administration of clonidine produces an increase in myocardial cGMP content by both a central and a peripheral action. The increase in cGMP can be due to a direct activation of cardiac prejunctional alpha-2 adrenoceptors and to stimulation of cardiac muscarinic receptors, a response secondary to an action of clonidine on central alpha-2 adrenoceptors.

Effect of halothane on myocardial cyclic AMP and cyclic GMP content of mice.

Halothane, in anesthetic concentrations (0.6-1.8 volumes/100 ml), produced a dose-dependent decrease in myocardial cyclic AMP (cAMP) content and an increase in cyclic GMP (cGMP) content in mice exposed to a continuous flow of the anesthetic carried in air for 15 min. Atropine (up to 20 mg/kg i.p.) did not alter significantly the myocardial cyclic nucleotides content or the effect of halothane on cAMP and cGMP content. Prazosin and yohimbine had no significant effect on cAMP or cGMP content in the absence of halothane. Both alpha adrenergic antagonists inhibited the halothane-induced increase in cGMP content (ID50, 0.24 and 0.54 mumol/kg i.p. for prazosin and yohimbine, respectively). In contrast, the decrease in cAMP content induced by halothane was not altered by alpha adrenergic antagonists. Propranolol (2 mg/kg i.p.) diminished myocardial cAMP level and prevented the halothane effect on myocardial cAMP content. Pretreatment with 6-hydroxydopamine did not change the cGMP response to halothane. Thus, the action of halothane on myocardial cyclic nucleotides content appears to be predominantly a peripheral effect, not related to cellular mechanisms mediated by muscarinic receptors. The results suggest that the increase in cGMP content induced by halothane does not require intact adrenergic nerve endings and that cellular processes associated with the alpha adrenoceptor system may be involved; the decrease in cAMP content may be due to an inhibition of the beta stimulatory action of catecholamines on adenylate cyclase.

Cardiac surgery causes desensitization of the beta-adrenergic receptor system of human lymphocytes.

Thirteen patients undergoing cardiac surgery were studied to examine whether beta-adrenergic desensitization occurs in the perioperative period surrounding cardiac surgery, using peripheral blood lymphocytes as a model. Lymphocytes were isolated before induction of anesthesia (PRE) and on the morning of the first postoperative day (POST). Cyclic adenosine monophosphate (cAMP) production from the lymphocytes was assayed in the untreated (BASAL) state, and after treatment with 5 microM isoproterenol, 10 microM prostaglandin E1, or 20 mM sodium fluoride with 10 microM AlCl3 (NaF). All cAMP values are reported as picomoles per 10(6) cells, mean +/- SEM. BASAL cAMP production did not change significantly between the PRE and POST samples (PRE, 1.2 +/- 0.1; POST, 1.0 +/- 0.1). Isoproterenol-stimulated cAMP was significantly lower postoperatively (PRE, 8.36 +/- 0.9; POST, 5.1 +/- 0.5; P less than 0.005). Prostaglandin E1-stimulated cAMP did not change (PRE, 21.7 +/- 2.4; POST, 25.3 +/- 2.5), and NaF-stimulated cAMP was increased postoperatively (PRE, 8.8 +/- 1.6; POST, 14.3 +/- 2.0; P less than 0.05). These findings suggest that cardiac surgery and/or cardiopulmonary bypass results in significant desensitization of the beta-adrenergic receptor/adenylate cyclase system of lymphocytes, which may parallel changes in the adrenergic response of other organ systems.