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Prostaglandin E1 (PGE1)-mediated transmembrane signal control systems were investigated in intact murine neuroblastoma cells (clone N1E-115). PGE1 increased intracellular levels of total inositol phosphates (IP), cyclic GMP, cyclic AMP, and calcium ([Ca2+]i). PGE1 transiently increased inositol 1,4,5-trisphosphate formation, peaking at 20 s. There was more than a 10-fold difference between the ED50 for PGE1 at cyclic AMP formation (70 nM) and its ED50 values at IP accumulation (1 microM), cyclic GMP formation (2 microM), and [Ca2+]i increase (5 microM). PGE1-mediated IP accumulation, cyclic GMP formation, and [Ca2+]i increase depended on both the concentration of PGE1 and extracellular calcium ions. PGE1 had more potent intrinsic activity in cyclic AMP formation, IP accumulation, and cyclic GMP formation than did PGE2, PGF2 alpha, or PGD2. A protein kinase C activator, 4 beta-phorbol 12 beta-myristate 13 alpha-acetate, had opposite effects on PGE1-mediated IP release and cyclic GMP formation (inhibitory) and cyclic AMP formation (stimulatory). These data suggest that there may be subtypes of the PGE1 receptor in this clone: a high-affinity receptor mediating cyclic AMP formation, and a low-affinity receptor mediating IP accumulation, cyclic GMP formation, and intracellular calcium mobilization.

The binding of [3H]neurotensin(8-13) to membranes from human frontal cortex at 0 degree C was time dependent, specific, saturable, and reversible. Saturation isotherms provided an equilibrium dissociation constant (KD) of 0.52 nM, and the maximal number of binding sites (Bmax) was 3.5 pmol/g original wet weight of tissue. Scatchard analysis yielded a straight line, and the Hill coefficient was equal to 1, a result indicating that [3H]neurotensin(8-13) bound to single, noncoopertive sites. The KD values of several analogs of neurotensin determined in competition with [3H]neurotensin(8-13) were similar to those previously determined in competition with [3H]neurotensin. The regional distribution of binding sites for [3H]neurotensin(8-13) was also similar to that for [3H]neurotensin. These results suggest that [3H]neurotensin(8-13) binds to the same sites as [3H]neurotensin and that [3H]neurotensin(8-13) has a higher affinity than [3H]neurotensin for these sites in human brain.

The binding of [3H]neurotensin to membranes from human brain at 0 degrees C was specific, saturable, and reversible. In the frontal cortex, the equilibrium dissociation constant (KD) for [3H]neurotensin determined from the ratio of rate constants (k-1/k1), saturation isotherms, and inhibition binding experiments was 0.80, 2.0, and 2.0 nM, respectively, and the maximum number of binding sites (Bmax) from the saturation isotherms and the competitive binding experiments was 2.4 and 2.2 pmol/g of tissue, respectively. Hill coefficients for binding were equal to 1, indicating the presence of single, noncooperative binding sites. Inhibition of specific binding of [3H]-neurotensin by several analogs of neurotensin showed that [Gln4]neurotensin and neurotensin(8-13) had the highest affinities for these binding sites in human frontal cortex, with each analog being approximately 13-fold more potent than neurotensin. In addition, these data showed that the carboxy-terminal portion of neurotensin played an important part in the binding of this neuropeptide in human brain, a result described for other species. Regional distribution of binding sites was different from that reported for animal brains. Of the 33 different regions investigated, the uncus and substantia nigra showed the highest specific binding of [3H]neurotensin, whereas such areas as the pineal body, medulla, and corpus callosum had few binding sites.