a G protein - dependent depolarizing action of sulphated cholecystokinin ocatapeptide ( CCK - 8 s ) on rodent neonatal spinal ventral horn neurons

Reports of cholecystokinin (CCK) binding and expression of CCK receptors in neonatal rodent spinal cord suggest that CCK may influence neuronal excitability. In patch-clamp recordings from 19/21 ventral horn motoneurons in neonatal (PN 5-12 days) rat spinal cord slices, we noted a slowly rising and prolonged membrane depolarization induced by bath applied sulfated CCK octapeptide (CCK-8s; 1 μM), blockable by the CCKB receptor antagonist L-365,260 (1 μM). Responses to non-sulfated CCK-8 or CCK-4 were significantly weaker. Under voltage clamp (VH −65mV), 22 / 24 motoneurons displayed a CCK-8s-induced tetrodotoxin-resistant inward current (peak −136 ± 28 pA) with a similar time course, mediated via reduction in a potassium conductance. In 29/31 unidentified neurons, CCK-8s induced a significantly smaller inward current (peak −42.8 ± 5.6 pA) and I-V plots revealed either membrane conductance decrease with net inward current reversal at −103.8 ± 4.7 mV (n=16), membrane conductance increase with net current reversing at −37.9 ± 3.6 mV (n=4), or parallel shift (n=9). Intracellular GTP-γ-S significantly prolonged the effect of CCK-8s (n=6), whereas GDP-β-S significantly reduced the CCK-8s response (n=6). Peak inward currents were significantly reduced after 5 min perfusion with N-ethylmaleimide. In isolated neonatal mouse spinal cord preparations, CCK-8s (30 nM300 nM) increased the amplitude and discharge of spontaneous depolarizations recorded from lumbosacral ventral roots. These observations imply functional postsynaptic G-protein coupled CCKB receptors are prevalent in neonatal rodent spinal cord. Page 2 of 34