Laboratorio de Fisiología y Biología Molecular, Instituto de Fisiología, Biología Molecular y Neurociencias (CONICET), Departamento de Fisiología y Biología Molecular y Celular, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires, Argentina.
PLoS One. 2013;8(1):e54846. doi: 10.1371/journal.pone.0054846. Epub 2013 Jan 30.
It is generally accepted that the immediately releasable pool is a group of readily releasable vesicles that are closely associated with voltage dependent Ca(2+) channels. We have previously shown that exocytosis of this pool is specifically coupled to P/Q Ca(2+) current. Accordingly, in the present work we found that the Ca(2+) current flowing through P/Q-type Ca(2+) channels is 8 times more effective at inducing exocytosis in response to short stimuli than the current carried by L-type channels. To investigate the mechanism that underlies the coupling between the immediately releasable pool and P/Q-type channels we transiently expressed in mouse chromaffin cells peptides corresponding to the synaptic protein interaction site of Cav2.2 to competitively uncouple P/Q-type channels from the secretory vesicle release complex. This treatment reduced the efficiency of Ca(2+) current to induce exocytosis to similar values as direct inhibition of P/Q-type channels via ω-agatoxin-IVA. In addition, the same treatment markedly reduced immediately releasable pool exocytosis, but did not affect the exocytosis provoked by sustained electric or high K(+) stimulation. Together, our results indicate that the synaptic protein interaction site is a crucial factor for the establishment of the functional coupling between immediately releasable pool vesicles and P/Q-type Ca(2+) channels.
普遍认为,即刻可释放池是一组与电压依赖性 Ca2+通道密切相关的易于释放的囊泡。我们之前已经表明,该池的胞吐作用与 P/Q Ca2+电流特异性偶联。因此,在本工作中我们发现,与 L 型通道携带的电流相比,通过 P/Q 型 Ca2+通道流动的 Ca2+电流在短刺激下引发胞吐作用的效率要高 8 倍。为了研究即刻可释放池与 P/Q 型通道之间偶联的机制,我们在小鼠嗜铬细胞中瞬时表达了与 Cav2.2 突触蛋白相互作用位点相对应的肽,以竞争性地将 P/Q 型通道与分泌囊泡释放复合物解偶联。这种处理将 Ca2+电流诱导胞吐作用的效率降低到与通过 ω-芋螺毒素-IVA 直接抑制 P/Q 型通道相似的值。此外,相同的处理显著降低了即刻可释放池的胞吐作用,但不影响持续电刺激或高 K+刺激引起的胞吐作用。总之,我们的结果表明,突触蛋白相互作用位点是即刻可释放池囊泡与 P/Q 型 Ca2+通道之间功能性偶联建立的关键因素。