Gérard Taton

Effects of PHI on vasoactive intestinal peptide receptors and adenylate cyclase activity in lung membranes. A comparison in man, rat, mouse and guinea pig

The presence of receptors, recognized by vasoactive intestinal peptide (VIP) as well as by PHI (a peptide with N-terminal histidine and C-terminal isoleucine amide), was documented in lung membranes from rat, mouse, guinea pig and man by the ability of these receptors, once occupied, to stimulate adenylate cyclase. In lung membranes from rat, mouse and guinea pig, the capacity of VIP, PHI and secretin to stimulate the enzyme and the potency of the same peptides to compete with 125I-VIP for binding to VIP receptors were similar, the affinity decreasing in the order: VIP greater than PHI greater than secretin. In addition, dose-effect curves were compatible with the coexistence of high-affinity and low-affinity VIP receptors, in the four animal species considered. If PHI was able to recognize all VIP receptors it could not, however, discriminate the subclasses of VIP receptors.

Vasoactive Intestinal Peptide (VIP) and peptide having N-terminal histidine and C-terminal isoleucine amide (PHI) stimulate adenylate cyclase activity in human heart membranes

The presence of receptors, recognized by Vasoactive Intestinal Peptide (VIP) and Peptide having N-terminal Histidine and C-terminal Isoleucine amide (PHI), was documented in membranes from human right auricle and left ventricular cardiac muscle by the ability of these peptides to stimulate adenylate cyclase. The capacity of VIP and PHI to activate the enzyme was comparable, in auricle as well as ventricle membranes, the affinity of the system being moderately higher for VIP than for PHI. In auricles, dose-effect curves appeared compatible with the coexistence of high-affinity and low-affinity VIP receptors. PHI could not, however, discriminate these subclasses of VIP receptors.

A comparison between muscarinic receptor occupancy, adenylate cyclase inhibition, and inotropic response in human heart.

SummaryBinding to muscarinic receptors was compared with adenylate cyclase inhibition in membranes derived from human heart auricles, and with inhibition of the contraction of auricular muscle fibers.In the absence of GTP, agonists recognized two classes of receptors both of which bound antagonists with the same affinity. In the presence of GTP, both classes of receptors for agonists were converted into a single low affinity state.Carbachol and oxotremorine inhibited adenylate cyclase activity by 43%, pilocarpine being less efficient (−28%). The 3 agonists exerted similar inhibitory effects on the inotropic response, in 7 out of 9 preparations of electrically- and norepinephrine-stimulated fibers. Dose-effect curves suggested that spareness (or an amplification mechanism) was implicated in the occupancy of low affinity binding sites by carbachol and oxotremorine (but not by the partial agonist pilocarpine) and the resulting inhibition of both adenylate cyclase activity and contractile force.

Characterization of the VIP- and secretin-stimulated adenylate cyclase system from human lung.

The response of a crude particulate adenylate cyclase preparation from surgically removed human lung to guanine nucleotides, sodium fluoride, β-adrenergic agonists, prostaglandins, vasoactive intestinal peptide (VIP), secretin, and [Val5]secretin was investigated. The enzyme activity increased 5, 10, and 9-fold, respectively, with GTP, Gpp(NH)p, and sodium fluoride. This activity was stimulated (in the presence as well as in the absence of added GTP) byd,l-isoproterenol,l-epinephrine andl-norepinephrine, the relative potency of these agonists being compatible with the existence of β-adrenoceptors of the β-adrenoceptors of the β2 subtype. Prostaglandins E1 and E2, but not PGF1α and PGF2α, stimulated the enzyme, PGE1 being at least 10 times more potent than PGE2. The biphasic pattern of stimulation of the same adenylate cyclase activity by VIP suggested the presence of high- and low-affinity VIP receptors coupled to the enzyme. This stimulation by VIP was not inhibited by secretin-(7–27). The stimulation of adenylate cyclase by secretin and [Val5]secretin was also biphasic, suggesting the coexistence of high- and low-affinity secretin receptors. Secretin-(7–27) was able to inhibit completely the secretin stimulation acting through high-affinity secretin receptors but exerted no effect on the stimulation operating through low-affinity secretin receptors, which might indicate that the latter receptors were in fact “VIP-preferring receptors”. [Val5]secretin was also used to differentiate these peptide receptors, since its properties were more VIP-like than those of secretin.

Comparison of VIP-secretin receptors in rat and human lung

The binding of 125I-VIP (Vasoactive Intestinal Peptide) to a crude particulate fraction from a rat lung was reversible and 125-I-VIP dissociation was accelerated by guanine triphosphate nucleotides. The relative potency of VIP and related peptides to compete with 125-VIP for binding was similar to their ability to stimulate adenylate cyclase in the same preparation. Dose-effect curves were compatible with the existence of two classes of VIP-receptors: a high-affinity type with equal affinity for VIP and [Val5] secretin, and a low-affinity type with affinity decreasing in the order VIP greater than [Val5] secretin greater than secretin. The response of a crude particulate adenylate cyclase preparation from human lung was also investigated. The biphasic pattern of adenylate cyclase stimulation by VIP suggested the presence of high- and low-affinity VIP receptors coupled to the enzyme. In addition, the stimulation of adenylate cyclase by secretin and [Val5] secretin was also biphasic, suggesting the coexistence of high- and low-affinity secretin receptors, Secretin (7-27) inhibited completely the secretin-stimulated activity operating through high-affinity secretion receptors, so that these receptors appear to be genuine secretin-preferring receptors.

Effects of full and partial β-adrenergic agonists and antagonists on human lung adenylate cyclase

The beta-adrenergic stimulation of adenylate cyclase in membranes from human lung was compared to that of adenylate cyclase in membranes with a majority of beta 2-adrenergic receptors (from rat lung) and in membranes with a homogeneous population of beta 2-adrenergic receptors (from rat erythrocytes and reticulocytes). In terms of adenylate cyclase stimulation, three full agonists (isoproterenol, epinephrine and norepinephrine), four partial agonists (procaterol, salbutamol, fenoterol and zinterol), and four antagonists (propranolol, metoprolol, atenolol and practolol) were tested. The potency (Kact or Ki) of the eleven beta-adrenergic agents, and the Hill coefficient (of 1) for the four antagonists tested indicated that the activation of human lung adenylate cyclase occurred through receptors of the beta 2-subtype only. Partial beta-adrenergic agonists were efficiently discriminated by the human lung preparation, as shown by distinct intrinsic activities. The mediocre efficacy and the relatively low potency of all beta-adrenergic agonists on adenylate cyclase suggested a relatively low density of beta 2-adrenergic receptors, as compared to the enzyme density.

Modulation by Indomethacin or Prostaglandin E2 of the Incidence of Diethylnitrosamine-induced γ-Glutamyltranspeptidase-positive Foci in Rat Liver

We investigated the effect of a pretreatment with indomethacin (IMC, ip 3.6 mg/kg body weight (bw)) or dimethylprostaglandin E2 (PGE2, ip 10 μg/kg bw) on the incidence and development of γ‐glutamyl‐transpeptidase (GGT)‐positive foci of altered hepatocytes, scored 8 or 14 weeks after ip injection of diethylnitrosamine (DENA, 50 mg/kg bw) to rats submitted to two‐thirds hepatectomy (PH) or sham operation (Sh). IMC reduced by about 4 times the incidence of DENA‐induced GGT‐positive foci per cm3 of liver tissue in sham‐operated as well as in two‐thirds hepatectomized rats, compared to the respective unpretreated controls. In contrast, PGE2 pretreatment increased the incidence of DENA‐induced foci in both groups, this effect, in terms of absolute numbers of foci, being additive to that of PH alone. IMC pretreatment resulted in foci with lower average size in the Sh but not in the PH animals, whereas with PGE2 pretreatment the mean volume of the foci was increased in the two groups of rats. At the dose used, IMC did not modify the proliferative response of hepatocytes to PH, and PGE2 did not stimulate proliferation in the sham‐operated animals. Altogether, these results indicate that: 1, the incidence of DENA‐induced foci can be negatively modulated by interfering with the prostaglandins pathway through a mechanism that does not involve an action either on proliferative activity or on any other process that would be specific to the post‐hepatectomy regenerative state; 2, positive modulation of the incidence of DENA‐induced foci does not necessarily require stimulation of proliferation.

Modulation of rat pancreatic muscarinic cholinergic receptors by caerulein.

To evaluate the modulation of pancreatic muscarinic receptors in two states of pancreatic growth, hypertrophy and hyperplasia, caerulein, a cholecystokinin analog, (1 microgram/kg) was administered thrice daily for 2 and 4 days to adult rats. After 2 days of treatment, pancreatic hypertrophy was well established as evidenced by increases in pancreatic weight, cellular mass and protein content. Using an increase in DNA content as an index of hyperplasia, we demonstrated that pancreatic hyperplasia occurred only after 4 days of caerulein treatment. Caerulein increased the concentration of muscarinic receptors per DNA in pancreatic homogenate by 57% over control value after 2 days of treatment without modification of the receptor affinity for the ligand QNB. This increase involved mainly receptors in the low affinity state for carbamylcholine and their concentration returned to control levels after 4 days of treatment. The functional capacity of the acini was significantly increased after 2 days of caerulein as amylase release (U/mg DNA) was significantly increased but the sensitivity of these acini to carbamylcholine was significantly decreased. After 4 days of caerulein, the functional capacity has returned towards control values but the sensitivity to carbamylcholine remained decreased. The increase in muscarinic receptor concentration could be ascribed to a general increase in cellular proteins, as part of the hypertrophic effect of caerulein. This specific effect would also explain the increased functional secretory capacity of the caerulein-treated acini but the decreased sensitivity to carbamylcholine probably resulted in changes at a postreceptor loci since the affinities of the muscarinic receptors for carbamylcholine remained unaffected.

Modulation by estrogen of the incidence of diethylnitrosamine-induced gamma-glutamyltranspeptidase-positive foci in rat liver

We measured the number and size of foci of altered hepatocytes induced after 8 weeks by diethylnitrosamine (DENA) in the liver of rats pretreated with 17 beta-estradiol (E2), 1 or 24 h prior to the administration of the carcinogen. The average size of the lesions was the same in the E2 pretreated and unpretreated animals. The number of gamma-glutamyltranspeptidase (GGT)-positive foci per cm3 of liver increased from 364 +/- 57 in unpretreated animals to 1149 +/- 186 in animals receiving E2 24 h before DENA; it raised to 3779 +/- 280 when the hormone was injected 1 h before the carcinogen, i.e. about 25% of the number of foci scored in rats receiving the carcinogen 24 h after partial hepatectomy. The hypothesis is proposed that 1-h pretreatment with E2 increases hepatocyte susceptibility towards DENA action by enhancing the accessibility of the genome to the carcinogen.