RESUMO
TREK-1 potassium channels are involved in a number of physiopathological processes such as neuroprotection, pain and depression. Molecules able to open or to block these channels can be clinically important. Having a cell model for screening such molecules is of particular interest. Here, we describe the development of the first available cell line that constituvely expresses the TREK-1 channel. The TREK-1 channel expressed by the h-TREK-1/HEK cell line has conserved all its modulation properties. It is opened by stretch, pH, polyunsaturated fatty acids and by the neuroprotective molecule, riluzole and it is blocked by spadin or fluoxetine. We also demonstrate that the h-TREK-1/HEK cell line is protected against ischemia by using the oxygen-glucose deprivation model.
Assuntos
Descoberta de Drogas/métodos , Avaliação Pré-Clínica de Medicamentos/métodos , Células HEK293/citologia , Doenças do Sistema Nervoso/tratamento farmacológico , Canais de Potássio de Domínios Poros em Tandem/metabolismo , Hipóxia Celular/efeitos dos fármacos , Ácidos Graxos Insaturados/farmacologia , Fluoxetina/farmacologia , Proteínas de Fluorescência Verde/metabolismo , Humanos , Concentração de Íons de Hidrogênio/efeitos dos fármacos , Ativação do Canal Iônico/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Técnicas de Patch-Clamp , Peptídeos/farmacologia , Canais de Potássio de Domínios Poros em Tandem/antagonistas & inibidores , Transporte Proteico/efeitos dos fármacos , Receptores de Superfície Celular/metabolismo , Riluzol/farmacologia , Estresse MecânicoRESUMO
We have characterized a cDNA encoding a Xenopus laevis apyrase (XAPY) that is expressed during embryogenesis. XAPY is highly homologous to two recently described mammalian apyrases, human SCAN-1 and rat Ca2+-NDPase, and to a lesser extent the salivary apyrase of the blood-feeding arthropod Cimex lectularis. RT-PCR analysis shows that Xapy is expressed at all the developmental stages tested, from oocytes through to tadpoles. Xapy transcripts are widely distributed in the embryo, but from late neurulae through to late tailbud stages they are highly enriched in the cement gland, an adhesive organ in the epidermis of the head. When expressed in HEK 293 cells, XAPY is largely retained in the endoplasmic reticulum, although some is also secreted. XAPY conditioned media hydrolyses UDP and UTP, confirming that it is a functional apyrase.