RESUMO
Neurotensin (NT) is a tridecapeptide distributed in central and peripheral nervous systems, which can behave as a neurotransmitter or neuromodulator at central and peripheral levels. Herein we tested the potential effect of this peptide on quinuclidinyl benzilate ([3H]-QNB) binding to muscarinic receptor in rat CNS membranes. It was observed that NT decreased up to 50-70% ligand binding at 1x10(-7) M-1x10(-5) M concentration in cerebral cortex, cerebellum and striatum. In the hippocampus, NT exerted a biphasic effect, behaving as a stimulator in the presence of 1x10(-12) M-1x10(-10) M concentration but as an inhibitor at 1x10(-8) M-1x10(-5) M concentration. In order to test the involvement of high-affinity NT receptor (NTS1) in NT inhibitory effect, assays were carried out in the presence of 1x10(-6) M NT and/or SR 48692 (Sanofi-Aventis, U.S., Inc.), a specific antagonist for this receptor, dissolved in dimethylsulfoxide (DMSO) 10% v/v. As controls, membranes incubated with DMSO and/or NT 1x10(-6) M plus DMSO were processed. It was found that NT+DMSO decreased [3H]-QNB binding to cerebral cortex, cerebellum and hippocampal membranes by 49%, 32% and 53%, respectively. This inhibition was not observed with the DMSO control group. Membrane preincubation with 1x10(-6) M SR 48692 failed to alter NT effect on binding. SR 48692 at 1x10(-6) M concentration decreased the binding by 50% only in cerebral cortex membranes, suggesting a possible direct effect of the antagonist on muscarinic receptors in this area. It was therefore concluded that the high-affinity NT receptor may not be involved in ligand binding inhibition to muscarinic receptor by NT.
Assuntos
Neurotensina/metabolismo , Receptores Muscarínicos/metabolismo , Receptores de Neurotensina/metabolismo , Animais , Masculino , Ratos , Ratos WistarRESUMO
Ontogenetic studies indicate that inositol phosphate accumulation in rodent brain tissue by cholinergic muscarinic agonists as well as expression of high-affinity neurotensin receptor (NTS1) peak at 7 days after birth. Herein, potential participation of this receptor in such effect was investigated. Cerebral cortex prisms of 7-day-old rats were preloaded with [3H]myoinositol and later incubated during 60 or 20 min in the presence of muscarinic agonist carbachol plus neurotensin and SR 48692, a non-peptide NTS1 antagonist. In 60-min incubation experiments, inositol phosphate accumulation by 10(-3) M carbachol was roughly 320%, an effect which remained unaltered plus 10(-6) M to 10(-4) M neurotensin but partially decreased with equimolar SR 48692 concentration. In 20-min incubation experiments, inositol phosphate accumulation by 10(-3) M carbachol was circa 240%, a value which attained 320-360% plus 10(-7) M neurotensin; this effect was totally blocked by 10(-7) M SR 48692. It was concluded that in inositol phosphate accumulation by carbachol, besides the cholinergic muscarinic receptor, the NTS1 receptor is likewise involved; findings at 60 min are attributable to the effect of endogenous neurotensin whereas those at 20 min most likely involve both endogenous and exogenously added peptide.
Assuntos
Encéfalo/efeitos dos fármacos , Carbacol/farmacologia , Agonistas Colinérgicos/farmacologia , Fosfatidilinositóis/metabolismo , Receptores de Neurotensina/fisiologia , Animais , Animais Recém-Nascidos , Encéfalo/metabolismo , Relação Dose-Resposta a Droga , Interações Medicamentosas , Feminino , Hidrólise/efeitos dos fármacos , Fosfatos de Inositol/farmacocinética , Masculino , Neurotensina/farmacologia , Pirazóis/farmacologia , Quinolinas/farmacologia , Ratos , Ratos Wistar , Receptores de Neurotensina/antagonistas & inibidores , Fatores de Tempo , TrítioRESUMO
Neurotensin (NT), a 13-amino acid peptide, is widely distributed in the brain and peripheral tissues of several mammalian species including man. In adult rat brain NT can bind to two distinct sites, one of high and the other of low affinity, corresponding to NT(1) and NT(2) receptor, respectively; structurally unrelated to these two, a third NT receptor (NT(3)) has been described. We have previously shown that Na(+), K(+)-ATPase is inhibited by NT when using ATP as substrate. In order to determine whether K(+)-stimulated dephosphorylation of this enzyme is involved, we tested NT effect by using p-nitrophenylphosphate, a non-natural substrate. K(+)-p-nitrophenylphosphatase activity was inhibited 42% by NT at 8.6 x 10(-6) M using an incubation medium containing 2 mM KCl but was unaffected in the presence of 5 or 20 mM KCl; however, with such KCl concentrations, NT was enabled to inhibit enzyme activity ( congruent with 35%) provided a suitable ATP:NaCl mixture (0.6:45.0 mM) was added. Mg(2+)-p-nitrophenylphosphatase activity remained unaltered at all conditions tested. Since SR 48692, a selective non-peptide NT(1) antagonist, abolished NT effect, involvement of NT(1) receptor in enzyme inhibition is suggested.
Assuntos
4-Nitrofenilfosfatase/antagonistas & inibidores , Neurotensina/farmacologia , Potássio/farmacologia , Receptores de Neurotensina/fisiologia , 4-Nitrofenilfosfatase/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Cinética , Magnésio/farmacologia , Masculino , Nitrofenóis/metabolismo , Compostos Organofosforados/metabolismo , Fosforilação , Pirazóis/farmacologia , Quinolinas/farmacologia , Ratos , Ratos Wistar , Receptores de Neurotensina/antagonistas & inibidores , Sinaptossomos/efeitos dos fármacos , Sinaptossomos/enzimologiaRESUMO
Neurotensin is a peptide present in mammalian CNS and peripheral tissues, which may play a major role in neurotransmission or neuromodulation, subserving diverse physiological functions. We studied the effect of added neurotensin on ATPase activities in synaptosomal membranes isolated from rat cerebral cortex. Neurotensin at 3 x 10(-8)-3 x 10(-6) M concentration decreased 20-44% Na+,K+-ATPase activity but failed to modify Mg2+-ATPase activity; lower neurotensin concentrations (3 x 10(-14)-3 x 10(-10) M) had no effect on enzyme activities. This inhibitory effect was abolished by neurotensin heating, by enzyme preincubation with neurotensin during periods exceeding 10 min, or by adding 1 x 10(-6) M SR 48692, a high affinity neurotensin receptor antagonist. Levocabastine, which blocks low affinity neurotensin receptor, failed to alter enzyme inhibition by the peptide. It is suggested that the sodium pump may be a target for neurotensin effects at neuronal level involving the participation of high affinity neurotensin receptor.
Assuntos
Neurotensina/fisiologia , Receptores de Neurotensina/fisiologia , ATPase Trocadora de Sódio-Potássio/antagonistas & inibidores , Animais , Córtex Cerebral/enzimologia , Córtex Cerebral/ultraestrutura , Calefação , Técnicas In Vitro , Membranas Intracelulares/enzimologia , Masculino , Neurotensina/farmacologia , Piperidinas/farmacologia , Ratos , Ratos Wistar , Receptores de Neurotensina/metabolismo , Sinaptossomos/enzimologiaRESUMO
The present investigation was undertaken to determine whether Ang-(1-7) is able to modify ATPase activities in membrane fractions prepared from several tissues. In the presence of 10(-6) M Ang-(1-7), total (Na , K+, Mg2+)-ATPase activity decreased 31% in rat atrium and 13% in sheep atrium but was unmodified in sheep liver, rat ventricle or crude brain membranes. In rat brain synaptosomal membranes, Ang-(1-7) at 10(-8) and 10(-7) M concentrations activated Na+, K+-ATPase 20 and 24%, respectively. Rat kidney Na+, K+-ATPase activity decreased roughly 40-70% with 10(-10)-10(-6) M Ang-(1-7)), but increased 22% with 10(-12) M peptide concentration, thus indicating a biphasic effect. Our findings showing that ATPase from several tissues responds differently to Ang-(1-7) are attributable to enzyme tissue specificity.
Assuntos
Angiotensina II/farmacologia , ATPase de Ca(2+) e Mg(2+)/metabolismo , Fragmentos de Peptídeos/farmacologia , ATPase Trocadora de Sódio-Potássio/metabolismo , Angiotensina I , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/enzimologia , Rim/efeitos dos fármacos , Rim/enzimologia , Fígado/efeitos dos fármacos , Fígado/enzimologia , Miocárdio/enzimologia , Ratos , Ratos Wistar , Ovinos , Sinaptossomos/efeitos dos fármacos , Sinaptossomos/enzimologiaAssuntos
4-Nitrofenilfosfatase/metabolismo , ATPase de Ca(2+) e Mg(2+)/metabolismo , Córtex Cerebral/enzimologia , Membranas Intracelulares/enzimologia , Neurotensina/farmacologia , ATPase Trocadora de Sódio-Potássio/metabolismo , Sinaptossomos/enzimologia , Trifosfato de Adenosina/farmacologia , Animais , Cinética , Ratos , Ratos Wistar , Cloreto de Sódio/farmacologiaRESUMO
Calcitonin is a hormone peptide produced by the thyroid gland, whose best described role is to prevent bone reabsorption. It also participates in other biological functions, even at central nervous system level. We studied the effect of added calcitonin on ATPase and acetylcholinesterase activities in synaptosomal membranes isolated from rat cerebral cortex. Calcitonin at 10(-7) - 10(-5)M concentration decreased 20-40% Na+, K(+)-ATPase and 15-25% K(+)-p-nitrophenylphosphatase activities, and at 10(-6)-10(-5)M reduced 20-30% Mg(2+)-p-nitrophenylphosphatase activity. However, this peptide failed to modify Mg(2+) - and Ca(2+)-ATPase or acetylcholinesterase activities. Results suggest that the sodium pump may be a target for calcitonin effects at neuronal level. Thus, calcitonin inhibition of sodium/potassium transport through synaptic membranes supports a regulatory role of this peptide on neurotransmission.