Activation of neurotensin receptors and purinoceptors in human colonic adenocarcinoma cells detected with the microphysiometer.
Activation of endogenous neurotensin (NT) receptors and P2-purinoceptors expressed by human colonic adenocarcinoma HT-29 cells increased extracellular acidification rates that were detected in the microphysiometer. NT (pGlu-Leu-Tyr-Glu-Asn-Lys-Pro-Arg-Arg-Pro-Tyr-Ile-Leu), NT[8-13] (Arg-Arg-Pro-Tyr-Ile-Leu), NT[9-13] (Arg-Pro-Tyr-Ile-Leu), and NT1 (N alpha methyl-Arg-Lys-Pro-Trp-Tle-Leu [Tle = tert-leucine]) were full agonists, whereas XL 775 (N-[N-[2-[3-[[6-amino-1-oxo-2-[[(phenylmethoxy)carbonyl]-amino]hex yl]amino]phenyl]-3-(4-hydroxyphenyl)-1-oxo-2-propenyl]-L-isoleucyl]-L-le ucine) was a partial agonist for activating NT receptors expressed by HT-29 cells. Desensitization induced by NT was rapid and monophasic with 85% of the initial response lost by a 30-s exposure. Once initiated, the rate and extent of desensitization were similar for different concentrations of a given agonist, for agonists of different potencies, and for agonists of different efficacies, which suggests that desensitization may be independent of receptor occupancy or agonist efficacy. Resensitization was a much slower process, requiring 60 min before the full agonist response to NT was recovered. ATP, via P2-purinoceptors, also activated cellular acidification rates in a concentration-dependent manner. ATP induced a biphasic desensitization of purinoceptors with a loss of ca. 50% of the initial stimulation detectable between 30 and 90 s of exposure to the agonist. Desensitization of NT receptors did not influence the activation of P2-purinoceptors by ATP, suggesting there was no heterologous desensitization between the two types of receptors. Superfusion with NT receptor agonists for 15 min at concentrations that did not elicit changes in extracellular acidification rates blocked, in a concentration-dependent manner, the agonist response induced by 100 nM NT. This may reflect sequestration of the receptor. These results suggest that the high agonist affinity state of NT receptors may modulate receptor sequestration, whereas activation of the low agonist affinity state may be linked to cellular metabolism. Comparison of our results with published data found differences as well as similarities of NT responses among three lines of HT-29 cells.
Richards
M
M
Marion Merrell Research Institute, Strasbourg, France.
van Giersbergen
P
P
Zimmermann
A
A
Lesur
B
B
Hoflack
J
J
D000255:Adenosine Triphosphate / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D019073:HT29 Cells; D006801:Humans; D009496:Neurotensin / Q000494:pharmacology; D018028:Receptors, Neurotensin / Q000032:analysis*; D018048:Receptors, Purinergic P2 / Q000032:analysis*; D013618:Tachyphylaxis
D018028:Receptors, Neurotensin; D018048:Receptors, Purinergic P2; D009496:Neurotensin; D000255:Adenosine Triphosphate