Neuronal type nicotinic acetylcholine receptors (nAchRs) have recently been identified in small-cell lung carcinoma. We here show that both nicotine and cytisine stimulate [3H]serotonin release in a dose-dependent manner; this effect is antagonized by a-bungarotoxin (aBgtx) and a-conotoxin MI (czctx). Nicotine and cytisine stimulate in vitro SCLC proliferation and this effect is completely antagonized by both aBgtx and aCtx. By PCR analysis, we demonstrate the presence in SCLC of both the a, and the & nAchR subunits mRNA. These data show that nAchRs play an important role in the biology of SCLC, and that algtx-sensitive receptors of the a7 subtype are crucially involved in both the secretagogue and mitogenic effects of nicotinic agonists.

Nerve growth factor (NGF) has antiproliferative and differentiating effects on adenomas of neuroendocrine origin. Cell lines derived from small-cell lung carcinoma (SCLC), a very aggressive neuroendocrine tumor, express NGF receptors. The role of NGF in the control of proliferation and progression of this carcinoma, however, has never been investigated. Chronic exposure of NCI-N-592 and GLC8 SCLC cell lines to NGF remarkably inhibited their proliferation rate both in vitro and in vivo, prevented their anchorage-independent clonal growth in soft agar, impaired their invasive capacity in vitro, and abolished their tumorigenic potential in nude mice. The proliferative response of SCLC cell lines to nicotine was also remarkably impaired by in vitro NGF treatment. Furthermore, NGF treatment activates in SCLC cell lines the expression and secretion of NGF. NGF thus reverts SCLC cell lines to a noninvasive, nontumorigenic phenotype that does not respond to nicotine and produces NGF.Small-cell lung cancer (SCLC) is a very aggressive human tumor representing about 25% of all lung cancers (1). Although our understanding of the biology of SCLC is expanding rapidly, there have been no recent major advances in the treatment of this tumor, and overall survival has not changed significantly since the late 1970s (1). SCLC has some phenotypical properties of a neuroendocrine tumor such as expression of L-dopa decarboxylase (1, 2), bombesin͞gastrin-releasing peptide (1, 3), neuron-specific enolase (1), and voltage-operated calcium channels of neuronal type (4-6). Proliferation of SCLC is controlled by autocrine loops sustained by the secretion of different neurohormones and growth factors such as bombesin, insulin-like growth factor I, bradykinin, neurotensin, cholecystokinin, and vasopressin (1). A mitogenic loop mediated by serotonin (5-HT) and facilitated by neuronal-type nicotinic receptors through stimulation of 5-HT release (7, 8) has been recently shown to be operative in SCLC cell lines (9, 10).Despite the observation that SCLC cell lines express (11) the tyrosine kinase trkA receptor for NGF (12)(13)(14), the role of this neurotrophin in the control of proliferation, invasiveness, and tumorigenic potential of this tumor has never been investigated. This issue seems of critical relevance as it is now emerging that NGF is an antiproliferative and differentiation factor for various tumors of neuroendocrine origin (15-21). In particular, NGF has been consistently reported to induce differentiation of the pituitary tumor cell line GH 3 (15) and the insulinoma cell line RINm5F (16,17) and to suppress cell growth and tumorigenicity of human prolactin-secreting adenomas (18,19). In addition, we found that an autocrine loop mediated by NGF is operative in both normal pituitary lactotroph cells (20) and slowly proliferating, nontumorigenic prolactinomas but not in tumorigenic prolactinoma cells (18,21). The impact of this mechanism is such that the breakdown of the NGF-mediated autocrine loop in nontumorigenic prolactinoma...

Voltage-operated calcium channels play crucial roles in stimulus-secretion coupling in pancreatic beta cells. A growing body of evidence indicates that these channels in beta cells are heterogeneous. In particular, not all the high-threshold calcium channels expressed belong to the best known L-type. In rat insulinoma cells, for example, L, N, and P/Q-type channels are present, while in human beta cells L-type and P/Q-type dominate. Where present, N-type and P/Q-type channels participate, alongside with the dominant L-type, in the control of sugar- or depolarization-induced hormone release. Distinct biophysical properties and selective modulation of the channel subtypes are likely to play important physiological roles. T-type channels are involved in beta cell apoptosis, while calcium channel autoantibodies recognizing high-threshold channels in beta cells, have been described both in neurological and diabetic patients. Subtype-selective calcium channel drugs have the potential for being beneficial in beta cell pathological states.

Electrophysiological measurements of cell capacitance (Cm) and biochemical assays of [3H] serotonin ([3H]5-hydroxytryptamine or [3H]5-HT) release were combined to study the control of secretion in rat insulinoma RINm5F cells. Depolarizing pulses produced Cm changes (DeltaCm), indicative of exocytosis, with the same voltage and Ca2+ dependency as the inward Ca2+ currents (ICa). Ba2+ was able to substitute for Ca2+ in stimulating exocytosis, but not endocytosis. However, both the relative potency and kinetics of Ca2+-versus Ba2+-triggered exocytosis differed significantly. 5-HT synthesis and uptake were demonstrated in RINm5F cells. This allowed the use of [3H]5-HT to study hormone release from cell populations. [3H]5-HT was released in a depolarization-, Ca2+- and time-dependent manner. Ba2+ also substituted for Ca2+ in depolarization-induced [3H]5-HT release. Thapsigargin, used to deplete Ca2+ stores, had no effects on Ca2+-triggered Cm increases, but Ca2+-triggered [3H]5-HT release was abolished. Ba2+-triggered [3H]5-HT release, however, was only slightly affected by Ca2+ store depletion. Ba2+ was found to act directly as a secretagogue of [3H]5-HT in intact cells, but not in Cm measurements of voltage-clamped cells, suggesting that cell depolarization is a prerequisite for this action.

Human small-cell lung carcinoma (SCLC) cells express neuronal-like voltage-operated calcium channels (VOCCs) and release mitogenic hormones such as serotonin (5-HT). Opioid peptides, on the other hand, have been shown to reduce SCLC cell proliferation by an effective autocrine pathway. Here we show that in GLC8 SCLC cells, only delta-opioid receptor subtype mRNA is expressed. Consistently, the selective delta-opioid agonist [D-Pen2-Pen5]-enkephalin (DPDPE), but not mu and kappa agonists, potently and dose-dependently inhibits high-threshold (HVA) VOCCs in these cells. As in peripheral neurons, this modulation is largely voltage-dependent, mediated by pertussis toxin (PTX)-sensitive G-proteins, cAMP-independent, and mainly affecting N-type VOCCs. With the same potency and selectivity, DPDPE also antagonizes the Ca(2+)-dependent release of [3H]serotonin ([3H]5-HT) from GLC8 cells. However, DPDPE inhibits not only the depolarization-induced release, but also the Ca(2+)-dependent secretion induced by thapsigargin or ionomycin. This suggests that besides inhibiting HVA VOCCs, opioids also exert a direct depressive action on the secretory apparatus in GLC8 cells. This latter effect also is mediated by a PTX-sensitive G-protein but, contrary to VOCC inhibition, it can be reversed by elevations of cAMP levels. These results show for the first time that opioids effectively depress both Ca2+ influx and Ca(2+)-dependent hormone release in SCLC cells by using multiple modulatory pathways. It can be speculated that the two mechanisms may contribute to the opioid antimitogenic action on lung neuroendocrine carcinoma cells.