Walker M W, Ewald D A, Perney T M, Miller R J
Department of Pharmacological and Physiological Sciences, University of Chicago, Illinois 60637.
J Neurosci. 1988 Jul;8(7):2438-46. doi: 10.1523/JNEUROSCI.08-07-02438.1988.
Using 125I-labeled neuropeptide Y (NPY) and peptide YY (PYY), we demonstrated the existence of specific receptors for these peptides on rat dorsal root ganglion (DRG) cells grown in primary culture. Scatchard analysis of membrane homogenates indicated that the peptides bound to 2 populations of sites, with approximate affinities of 0.08 and 6.5 nM. Only low levels of binding were detected on sympathetic neurons cultured from the same animals or on a variety of neuronal clonal cell lines. The binding of 125I-NPY and 125I-PYY to DRG cell membranes was considerably reduced by the nonhydrolyzable analog of GTP, Gpp(NH)p. The major effect of Gpp(NH)p was to reduce the number of lower-affinity NPY binding sites without altering the number of high-affinity binding sites. NPY potently inhibited Ca2+ currents recorded under voltage clamp in rat DRG cells. Both the transient and sustained portions of the Ca2+ current were inhibited. The inhibitory effects of NPY were completely blocked following treatment of the cells with pertussis toxin. Depolarization elicited a large influx of Ca2+ into DRG neurons as assessed using fura-2-based microspectrofluorimetry. This influx of Ca2+ could be partially inhibited by NPY. Furthermore, NPY effectively inhibited the depolarization-induced release of substance P from DRG cells in vitro. Thus, NPY may be an important regulator of sensory neuron function in vivo.
我们使用125I标记的神经肽Y(NPY)和肽YY(PYY),证明了在原代培养的大鼠背根神经节(DRG)细胞上存在这些肽的特异性受体。对膜匀浆进行Scatchard分析表明,这些肽与两类位点结合,其近似亲和力分别为0.08和6.5 nM。在从同一动物培养的交感神经元或多种神经元克隆细胞系上仅检测到低水平的结合。不可水解的GTP类似物Gpp(NH)p可显著降低125I-NPY和125I-PYY与DRG细胞膜的结合。Gpp(NH)p的主要作用是减少低亲和力NPY结合位点的数量,而不改变高亲和力结合位点的数量。NPY能有效抑制大鼠DRG细胞电压钳记录下的Ca2+电流。Ca2+电流的瞬时和持续部分均受到抑制。用百日咳毒素处理细胞后,NPY的抑制作用被完全阻断。使用基于fura-2的显微分光荧光测定法评估,去极化可引起大量Ca2+流入DRG神经元。这种Ca2+内流可被NPY部分抑制。此外,NPY在体外能有效抑制去极化诱导的DRG细胞中P物质的释放。因此,NPY可能是体内感觉神经元功能的重要调节因子。