University of Toledo College of Medicine, Department of Neurosciences, Toledo, OH 43614-5804, USA.
Mol Cell Neurosci. 2010 Feb;43(2):244-57. doi: 10.1016/j.mcn.2009.11.007. Epub 2009 Dec 1.
Neuropeptides collaborate with conventional neurotransmitters to regulate synaptic output. Pituitary adenylate cyclase-activating polypeptide (PACAP) co-localizes with acetylcholine in presynaptic nerve terminals, is released by stimulation, and enhances nicotinic acetylcholine receptor- (nAChR-) mediated responses. Such findings implicate PACAP in modulating nicotinic neurotransmission, but relevant synaptic mechanisms have not been explored. We show here that PACAP acts via selective high-affinity G-protein coupled receptors (PAC(1)Rs) to enhance transmission at nicotinic synapses on parasympathetic ciliary ganglion (CG) neurons by rapidly and persistently increasing the frequency and amplitude of spontaneous, impulse-dependent nicotinic excitatory postsynaptic currents (sEPSCs). Of the canonical adenylate cyclase (AC) and phospholipase-C (PLC) transduction cascades stimulated by PACAP/PAC(1)R signaling, only AC-generated signals are critical for synaptic modulation since the increases in sEPSC frequency and amplitude were mimicked by 8-Bromo-cAMP, blocked by inhibiting AC or cAMP-dependent protein kinase (PKA), and unaffected by inhibiting PLC. Despite its ability to increase agonist-induced nAChR currents, PACAP failed to influence nAChR-mediated impulse-independent miniature EPSC amplitudes (quantal size). Instead, evoked transmission assays reveal that PACAP/PAC(1)R signaling increased quantal content, indicating that it modulates synaptic function by increasing vesicular ACh release from presynaptic terminals. Lastly, signals generated by the retrograde messenger, nitric oxide- (NO-) are critical for the synaptic modulation since the PACAP-induced increases in spontaneous EPSC frequency, amplitude and quantal content were mimicked by NO donor and absent after inhibiting NO synthase (NOS). These results indicate that PACAP/PAC(1)R activation recruits AC-dependent signaling that stimulates NOS to increase NO production and control presynaptic transmitter output at neuronal nicotinic synapses.
神经肽与传统神经递质协同调节突触输出。垂体腺苷酸环化酶激活肽（PACAP）与乙酰胆碱在突触前神经末梢共存，受刺激释放，并增强烟碱型乙酰胆碱受体（nAChR）介导的反应。这些发现表明 PACAP 参与调节烟碱能神经传递，但相关的突触机制尚未被探索。我们在这里表明，PACAP 通过选择性高亲和力 G 蛋白偶联受体（PAC1Rs）作用，通过快速而持久地增加自发性、脉冲依赖性烟碱兴奋性突触后电流（sEPSC）的频率和幅度，增强副交感睫状神经节（CG）神经元上的烟碱突触传递。在 PACAP/PAC1R 信号刺激的经典腺苷酸环化酶（AC）和磷脂酶 C（PLC）转导级联中，只有 AC 产生的信号对突触调节至关重要，因为 sEPSC 频率和幅度的增加被 8-溴-cAMP 模拟，被抑制 AC 或 cAMP 依赖性蛋白激酶（PKA）阻断，不受 PLC 抑制的影响。尽管 PACAP 能够增加激动剂诱导的 nAChR 电流，但它未能影响 nAChR 介导的脉冲非依赖性微小 EPSC 幅度（量子大小）。相反，诱发传递测定表明，PACAP/PAC1R 信号增加了量子含量，表明它通过增加来自突触前末端的囊泡 ACh 释放来调节突触功能。最后，逆行信使一氧化氮（NO）产生的信号对于突触调节至关重要，因为 PACAP 诱导的 sEPSC 频率、幅度和量子含量的增加被 NO 供体模拟，并且在抑制一氧化氮合酶（NOS）后消失。这些结果表明，PACAP/PAC1R 激活募集 AC 依赖性信号，刺激 NOS 增加 NO 产生并控制神经元烟碱能突触的前突递质输出。
