We suggest that Galpha9 functions in an inhibitory-feedback pathway that regulates cAMP signaling center formation and propagation. Galpha9 may be part of the mechanism that regulates lateral signal inhibition or that modulates receptor desensitization.

UC11 cells, derived from a human astrocytoma, have a high density of functional substance P receptors. Radioligand binding studies were conducted with the highly selective neurokinin-1 receptor ligand [3H][Sar9,Met(O2)11]-substance P. Kinetic binding experiments conducted at 4 degrees C yielded an association rate constant k1 of 1.86 x 10(7) M-1 min-1, a dissociation rate constant k-1 of 0.00478 min-1, and a calculated kinetic KD of 257 pM. Saturation binding experiments yielded average values of KD = 447 +/- 103 pM, Bmax = 862 +/- 93 fmol/mg of protein. This Bmax corresponds to more than 150,000 binding sites/cell. Competition binding experiments with unlabeled [Sar9,Met(O2)11]-substance P yielded average values of KD = 491 +/- 48 pM and Bmax = 912 +/- 67 fmol/mg of protein. In [3H]inositol-labeled cells, substance P induced a robust inositol phosphate formation. Inositol trisphosphate levels increased as much as 20-fold within approximately 15 s of addition of substance P. This inositol trisphosphate formation was transient and had returned to baseline within the first 60-120 s. Inositol monophosphate formation, however, was linear for at least 2 h. Structure activity data on binding and inositol monophosphate formation confirmed the presence of a neurokinin-1 receptor subtype in these cells. Thus, the UC11 cell should be a useful model cell for delineating the physiological role of substance P receptors in astrocytes.

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Histamine stimulated inositol phosphate formation by human skin fibroblasts. The effect of histamine was reduced but still readily apparent in the absence of extracellular Ca2+. Histamine caused a transient increase in intracellular free Ca2+ as detected by indo-1 and fura-2 fluorescence studies on cell populations and on individual cells. Similar increases were observed in the absence of extracellular Ca2+, indicating that the effect was primarily due to mobilization of Ca2+ from intracellular stores, presumably by inositol trisphosphate (IP3). The effects of histamine on phosphoinositide metabolism and intracellular Ca2+ were inhibited by pretreatment of the cells with phorbol esters, suggesting that the histamine receptor in fibroblasts is subject to feedback regulation by protein kinase C. Histamine inhibited the incorporation of [3H]-thymidine into DNA. The effects of histamine on inositol phosphate formation, intracellular Ca2+, and thymidine incorporation were blocked by the H1 receptor antagonist mepyramine. Our results indicate that human skin fibroblasts have H1 receptors coupled to the formation of inositol phosphates and mobilization of intracellular Ca2+. We suggest that this H1 receptor also mediates a block of the cell cycle and that histamine may play a physiological role in the regulation of fibroblast proliferation.