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1.
J Med Chem ; 43(9): 1741-53, 2000 May 04.
Artigo em Inglês | MEDLINE | ID: mdl-10794691

RESUMO

A set of novel tachykinin-like peptides has been isolated from bullfrog brain and gut. These compounds, ranatachykinin A (RTKA), ranatachykinin B (RTKB), and ranatachykinin C (RTKC), were named for their source, Rana catesbeiana, and their homology to the tachykinin peptide family. We present the first report of the micelle-bound structures and pharmacological actions of the RTKs. Generation of three-dimensional structures of the RTKs in a membrane-model environment using (1)H NMR chemical shift assignments, two-dimensional NMR techniques, and molecular dynamics and simulated annealing procedures allowed for the determination of possible prebinding ligand conformations. RTKA, RTKB, and RTKC were determined to be helical from the midregion to the C-terminus (residues 4-10), with a large degree of flexibility in the N-terminus and minor dynamic fraying at the end of the C-terminus. The pharmacological effects of the RTKs were studied by measuring the elevation of intracellular Ca(2+) in Chinese hamster ovarian cells stably transfected with the bullfrog substance P receptor (bfSPR). All of the RTKs tested elicited Ca(2+) elevations with a rank order of maximal effect of RTKA >/= SP > RTKC >/= RTKB. A high concentration (1 microM) of the neuropeptides produced varying degrees of desensitization to a subsequent challenge with the same or different peptide, while a low concentration (1 pM) produced sensitization at the bfSPR. Our data suggest differences in amino acid side chains and their charged states at the C-terminal sequence or differences in secondary structure at the N-terminus, which do not overlap according to the findings in this paper, may explain the differing degree and type of receptor activation seen at the bfSPR.


Assuntos
Receptores da Neurocinina-1/metabolismo , Taquicininas/metabolismo , Sequência de Aminoácidos , Animais , Células CHO , Cálcio/metabolismo , Canais de Cálcio/metabolismo , Cricetinae , Corantes Fluorescentes , Espectroscopia de Ressonância Magnética , Micelas , Modelos Moleculares , Dados de Sequência Molecular , Conformação Proteica , Rana catesbeiana , Receptores da Neurocinina-1/química , Dodecilsulfato de Sódio
2.
Peptides ; 24(3): 469-75, 2003 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-12732347

RESUMO

The actions of four tachykinins on inhibition and desensitization of the M-current of bullfrog sympathetic neurons have been characterized. Radioligand binding parameters of the tachykinins were determined at a neurokinin receptor in a heterologous expression system. The correlation between binding, signaling and receptor regulation was investigated. A correlation between receptor binding and signaling was found between the peptides; however, their ability to produce desensitization was not correlated with binding and signaling. These results show that the ability of a tachykinin peptide to induce signal activation is not indicative of its ability to induce receptor regulation.


Assuntos
Regulação para Baixo/efeitos dos fármacos , Receptores da Neurocinina-1/agonistas , Receptores da Neurocinina-1/metabolismo , Taquicininas/farmacologia , Sequência de Aminoácidos , Animais , Células CHO , Cricetinae , Eletrofisiologia , Ligantes , Potássio/metabolismo , Ligação Proteica , Rana catesbeiana , Taquicininas/química
3.
Genes Brain Behav ; 13(7): 579-91, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25077934

RESUMO

Maternal care is an indispensable component of offspring survival and development in all mammals and necessary for reproductive success. Although brain areas regulating maternal behaviors are innervated by serotonergic afferents, very little is known about the role of this neurotransmitter in these behaviors. To evaluate the contribution of serotonin to maternal care, we used mice with a null mutation in the gene for tryptophan hydroxylase-2 (TPH2), which results in a genetic depletion of brain serotonin, and tested them in a wide range of maternal behavior paradigms. We found that litters born to and reared by TPH2(-/-) mothers showed decreased survival, lower weaning weights and increased cannibalization. In addition, TPH2(-/-) mothers performed poorly in pup retrieval, huddling, nest construction and high-arched back nursing. Aggression in TPH2(-/-) dams was not triggered by lactation and was steadily high. Survival and weaning weight deficits of TPH2(-/-) pups were rescued by cross-fostering and in litters of mixed genotype (TPH2(-/-) and TPH2(-/+) ). However, the maternal behaviors of TPH2(-/-) dams did not improve when rearing either TPH2(+/+) pups or mixed-genotype litters. In addition, TPH2(-/-) pups significantly worsened the behavior of TPH2(+/+) dams with respect to cannibalism, weaning weight and latency to attack. Olfactory and auditory functions of TPH2(-/-) females or anxiety-like behaviors did not account for these maternal alterations as they were equal to their TPH2(+/+) counterparts. These findings illustrate a profound influence of brain serotonin on virtually all elements of maternal behavior and establish that TPH2(-/-) pups can engender maladaptive mothering in dams of both genotypes.


Assuntos
Encéfalo/metabolismo , Comportamento Materno , Serotonina/metabolismo , Triptofano Hidroxilase/metabolismo , Animais , Animais Recém-Nascidos/fisiologia , Feminino , Genótipo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Magreza , Triptofano Hidroxilase/genética
4.
J Pharmacol Exp Ther ; 313(3): 1347-54, 2005 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-15764733

RESUMO

Following agonist binding, neurokinin-1 receptors undergo rapid desensitization followed by internalization and recycling. Desensitization requires receptor phosphorylation but does not require internalization, whereas resensitization is thought to require internalization and recycling. Our previous data, however, have suggested that, following activation and desensitization, the return of responsiveness to the neurokinin-1 agonist substance P (termed "resensitization") occurs hours before internalized receptors are recycled back to the plasma membrane. To further investigate this novel mechanism of neurokinin-1 receptor resensitization, we have studied the time courses of neurokinin-1 receptor responsiveness, recycling, and dephosphorylation by measuring cellular Ca(2+) responses, ligand-receptor binding, and receptor phosphorylation, respectively. Concentration-response curves and competition binding curves were obtained at various times following desensitization. The effects of the nonhydrolyzable GTP analog Gpp(NH)p on substance P binding were also studied to assess receptor-G protein coupling. After receptor activation and desensitization, Ca(2+) signaling in response to substance P occurred within 90 min, whereas the return of receptor binding required 240 min. Receptor dephosphorylation was greater than 90% complete 20 min after agonist washout. In addition, the return of substance P responsiveness coincided with a return in sensitivity of substance P binding to Gpp(NH)p, indicating a return in receptor-G protein coupling. These data show that the resensitization of responsiveness to substance P precedes receptor recycling. This may result from a conversion of nonfunctional neurokinin-1 receptors to functional receptors at the plasma membrane.


Assuntos
Receptores da Neurocinina-1/metabolismo , Trifosfato de Adenosina/farmacologia , Animais , Ligação Competitiva , Células CHO , Cálcio/metabolismo , Cricetinae , Guanilil Imidodifosfato/farmacologia , Fosforilação , Ratos , Substância P/metabolismo
5.
J Pharmacol Exp Ther ; 303(3): 1155-62, 2002 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-12438539

RESUMO

Prolonged or repeated activation of many G protein-coupled receptors induces rapid desensitization followed by a period during which receptors are resensitized. In this study, concanavalin A (Con A) and monensin were used to investigate the mechanisms of desensitization and resensitization of the neurokinin-1 receptor. Con A inhibits internalization, whereas monensin prevents receptor recycling. The effects of Con A and monensin on desensitization, resensitization, receptor phosphorylation, endocytosis, and recycling of the neurokinin-1 receptor were assessed. Desensitization was defined as the decrease in the ability of substance P (SP) to elicit an intracellular Ca2+ response after a prolonged SP exposure. Resensitization was characterized as the return of SP responsiveness. Under control conditions, desensitization occurred after a 5-min exposure to agonist. Resensitization was evident 30 min after agonist washout. Neither monensin nor Con A prevented desensitization. Monensin completely inhibited resensitization, whereas Con A decreased but did not completely block resensitization. Receptor phosphorylation was increased after agonist activation and returned to basal levels after a recovery period. Neither Con A nor monensin altered the amount of agonist-specific receptor phosphorylation. Receptor binding analysis showed that plasma membrane receptors were internalized after a 5-min agonist exposure. Receptor recycling was not observed after a 1-h recovery period; however, resensitization was apparent. Taken together, these results suggest that rapid neurokinin-1 receptor desensitization can occur without receptor internalization and that resensitization occurs before receptor recycling.


Assuntos
Receptores da Neurocinina-1/agonistas , Receptores da Neurocinina-1/metabolismo , Animais , Células CHO , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Concanavalina A/farmacologia , Cricetinae , Monensin/farmacologia , Fosforilação/efeitos dos fármacos , Ratos
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