Uptake of norepinephrine and related catecholamines by cultured chromaffin cells: characterization of cocaine-sensitive and -insensitive plasma membrane transport sites.
(17/21)
Norepinephrine and its closely related analogues, dopamine and epinephrine, are transported into chromaffin cells in culture by two distinct types of sites on the plasma membrane: one is sensitive to cocaine while the other is not. The cocaine-sensitive site has a high affinity for catecholamines and depends on sodium in the medium. The apparent Km for norepinephrine uptake by the cocaine-sensitive site is 5.8 microM when determined in the presence of 118 mM NaCl, obtained using nonlinear least-square curve fitting. Detailed kinetic analysis has also shown cocaine to be a competitive inhibitor of norepinephrine uptake with an apparent Ki of ca. 1 microM. This site is blocked by a series of tricyclic antidepressant drugs with relative potencies characteristic of norepinephrine transport sites in neurons. In contrast, the cocaine-insensitive site(s) have a low affinity for norepinephrine (apparent Km, approximately 88 microM) and are also able to transport catecholamine analogues such as dimethyl-epinephrine and isoproterenol, which have bulky groups attached to the amine moiety. Transport of norepinephrine at both sites is blocked by low temperature, by mitochondrial uncouplers, and by other metabolic inhibitors. Both of these transport sites in the chromaffin cell plasma membrane, therefore, appear to be different from the well-characterized catecholamine transport sites in the chromaffin granule membrane on the basis of substrate specificity and their sensitivity to inhibitors. (+info)
Behavioral and neurochemical changes produced by postnatal pretreatments with methamphetamine in rats.
(18/21)
Male neonates of Wistar strain rats were given s.c. 1-4 mg/kg/day of methamphetamine (MAP) for 7 successive days from days 6 to 12 after birth. The acquisition processes of the discriminative lever-press avoidance response were investigated from day 60 after birth. Effects of the postnatal pretreatments with MAP on saturation constants for specific bindings of 3H-spiperone (SPP) and 3H-WB4101, respectively, in 8 brain regions were also investigated at 100-120 days after birth. In addition, dopamine, noradrenaline and the levels of their metabolites were measured in the brain. No significant difference was detected in body weight, gross behaviors and avoidance learning between saline- and MAP-pretreated groups. However, effects of MAP and apomorphine on the locomotor activity significantly increased in the MAP-pretreated group. Significant decreases in Bmax and Kd values of 3H-SPP binding sites in the striatum were detected in the MAP-pretreated group, while significant decreases in Bmax values of 3H-WB4101 binding sites in the cortex and hippocampus as well as those in Kd values in the hippocampus were found in the treated group. Dopamine and noradrenaline levels significantly decreased in the MAP-pretreated group, but on the contrary, their metabolites levels significantly increased. These results suggest that postnatal pretreatments with MAP produce long-lasting behavioral changes associated with an impaired development of catecholaminergic neurons in the rat brain after maturity. (+info)
Effects of stimulation of muscarinic and of beta-catecholamine receptors on the intracellular distribution of protein kinase C in guinea pig exocrine glands.
(19/21)
Stimulation of exocrine cells via muscarinic receptors is associated with an activation of protein kinase C [Padel & Soling (1985) Eur. J. Biochem. 151, 1-10]. We show here that stimulation of isolated parotid gland lobules with 8 X 10(-6) M-carbamoylcholine leads to a translocation of protein kinase C from the cytosolic to the particulate compartment within 30 s (25% and 45% of total activity recovered in the particulate fraction of controls and stimulated samples respectively). The specific enzyme activity in the particulate fraction increased to 169% of the corresponding control value. After 10 min the changes started to reverse and, after 30 min, cytosolic protein kinase C was higher in stimulated than in unstimulated lobules. Isoproterenol (2 X 10(-5) M) stimulated the release of amylase more than did carbamoylcholine, but did not significantly affect intracellular distribution of protein kinase C during the observation time of 30 min. In isolated pancreatic lobules a significant carbamoylcholine-mediated translocation of protein kinase C into the particulate fraction could be observed after 5 and 20 min, but not after 1 min. After 5 min the specific enzyme activity in the particulate fraction had increased to 153% of the corresponding controls. The corresponding decrease (-38%) in the specific enzymic activity of cytosolic protein kinase C stayed constant up to 30 min. In isolated parotid gland lobules alpha-amylase secretion proceeded at a linear rate already during the first 1 min of stimulation, whereas in pancreatic lobules a measurable rate of alpha-amylase secretion did not occur before 5 min. These differences in time course paralleled the differences in the onset of translocation of protein kinase C. The results support a direct involvement of protein kinase C in carbamoylcholine-mediated but not in isoproterenol-mediated stimulation of exocytosis in exocrine cells. (+info)
Molecular cloning of a fish gene encoding a novel seven-transmembrane receptor related distantly to catecholamine, histamine, and serotonin receptors.
(20/21)
A genomic DNA fragment encoding a G protein-coupled seven-transmembrane receptor was isolated from Medaka fish, Oryzias latipes. The encoded protein is similar in sequence to other receptors including catecholamine, histamine and serotonin receptors. However, the similarity is much lower than those among members of these receptor subfamilies, thus suggesting this seven-transmembrane receptor to be an orphan receptor whose ligand has not yet been identified. Genomic Southern blot analysis suggested that the fish genome contains additional receptor genes related to the isolated gene, indicating that this novel receptor, possibly with its related receptors, might constitute a novel subfamily of the seven-transmembrane receptor superfamily. (+info)
Cardiac-specific overexpression of angiotensin II AT2 receptor causes attenuated response to AT1 receptor-mediated pressor and chronotropic effects.
(21/21)
Angiotensin (Ang) II has two major receptor isoforms, AT1 and AT2. Currently, AT1 antagonists are undergoing clinical trials in patients with cardiovascular diseases. Treatment with AT1 antagonists causes elevation of plasma Ang II which selectively binds to AT2 and exerts as yet undefined effects. Cardiac AT2 level is low in adult hearts, whereas its distribution ratio is increased during cardiac remodeling and its action is enhanced by application of AT1 antagonists. Although in AT2 knock-out mice sensitivity to the pressor action of Ang II was increased, underlying mechanisms remain undefined. Here, we report the unexpected finding that cardiac-specific overexpression of the AT2 gene using alpha-myosin heavy chain promoter resulted in decreased sensitivity to AT1-mediated pressor and chronotropic actions. AT2 protein undetectable in the hearts of wild-type mice was overexpressed in atria and ventricles of the AT2 transgenic (TG) mice and the proportions of AT2 relative to AT1 were 41% in atria and 45% in ventricles. No obvious morphological change was observed in the myocardium and there was no significant difference in cardiac development or heart to body weight ratio between wild-type and TG mice. Infusion of Ang II to AT2 TG mice caused a significantly attenuated increase in blood pressure response and the change was completely blocked by pretreatment with AT2 antagonist. This decreased sensitivity to Ang II-induced pressor action was mainly due to the AT2-mediated strong negative chronotropic effect and exerted by circulating Ang II in a physiological range that did not stimulate catecholamine release. Isolated hearts of AT2 transgenic mice perfused using a Langendorff apparatus also showed decreased chronotropic responses to Ang II with no effects on left ventricular dp/dt max values, and Ang II-induced activity of mitogen-activated protein kinase was inhibited in left ventricles in the transgenic mice. Although transient outward K+ current recorded in cardiomyocytes from AT2 TG mice was not influenced by AT2 activation, this study suggested that overexpression of AT2 decreases the sensitivity of pacemaker cells to Ang II. Our results demonstrate that stimulation of cardia AT2 exerts a novel antipressor action by inhibiting AT1-mediated chronotropic effects, and that application of AT1 antagonists to patients with cardiovascular diseases has beneficial pharmacotherapeutic effects of stimulating cardiac AT2. (+info)