RESUMEN
Two small conductance calcium-activated potassium channels that exhibited amino acid similarity with mammalian SK2 and SK3, respectively, were PCR cloned from the CNS of trout and sequenced. Upon heterologous expression in Sf21 insect cells trout SK2 (termed tSK2) produced a calcium-dependent, voltage-insensitive, and non-inactivating current with a single unit conductance of about 11 pS. This current was half maximally activated by 0.76 microM Ca(2+) and blocked by picomolar concentrations of apamin but not by TEA. Transcripts of both SK-related channels possessed a widespread distribution in the mature brain tissue of trout and outside the nervous system were detectable in muscle tissue as well as in liver. As revealed by RT-PCR analysis transcripts encoding tSK2 and tSK3 channels were early detectable during brain development (stage 30, shortly after hatching).
Asunto(s)
Membrana Celular/metabolismo , Sistema Nervioso Central/metabolismo , Larva/metabolismo , Neuronas/metabolismo , Oncorhynchus mykiss/metabolismo , Canales de Potasio Calcio-Activados , Canales de Potasio/genética , Canales de Potasio/aislamiento & purificación , Animales , Calcio/metabolismo , Calcio/farmacología , Membrana Celular/efectos de los fármacos , Sistema Nervioso Central/citología , Sistema Nervioso Central/crecimiento & desarrollo , ADN Complementario/genética , ADN Complementario/aislamiento & purificación , Evolución Molecular , Regulación del Desarrollo de la Expresión Génica/fisiología , Larva/anatomía & histología , Larva/crecimiento & desarrollo , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/genética , Datos de Secuencia Molecular , Estructura Molecular , Neuronas/efectos de los fármacos , Oncorhynchus mykiss/anatomía & histología , Oncorhynchus mykiss/crecimiento & desarrollo , Filogenia , Bloqueadores de los Canales de Potasio/farmacología , Canales de Potasio/efectos de los fármacos , Homología de Secuencia de Aminoácido , Homología de Secuencia de Ácido Nucleico , Canales de Potasio de Pequeña Conductancia Activados por el CalcioRESUMEN
Two Shaw-related potassium channels expressed in the CNS of trout were PCR cloned and sequenced: Traw1 was identified as a fish homologue to mammalian K(v)3.1, while Traw2 could not be exactly classified. Upon heterologous expression Traw1 exhibited a high threshold (-20 mV) non-inactivating delayed rectifier current that was efficiently blocked by submicromolar concentrations of TEA, 4-AP and quinine but not by alpha-DTX or apamin. The amplitude of the Traw1 induced current was reduced by the phorbol ester TPA, the effect being prevented by the proteinkinase inhibitor H7. Transcripts of both Shaw- related channels possess a widespread distribution in the mature brain tissue and outside the nervous system are detectable in muscle but not in liver. During brain development Traw1 mRNA was initially identified at stage 31 (shortly after hatching) while transcripts encoding Traw2 were detectable already at stage 28 (1 week before hatching).