Guinea-pig Receptors Research Articles

P-242 - Endothelin-1 induced bronchoconstriction was suppressed by only blockade of both ETA and ETB receptors in guinea pigs and humans.

P-242 - Endothelin-1 induced bronchoconstriction was suppressed by only blockade of both ETA and ETB receptors in guinea pigs and humans.

Binding of spironolactone metabolites in vivo to renal mineralocorticoid receptors in guinea pigs.

Spironolactone (SL) is a mineralocorticoid antagonist used clinically to treat hypertension and congestive heart failure. Its mechanism of action involves competitive binding to aldosterone receptors in the kidneys, resulting in diuresis. It is known that the actions of SL are mediated by metabolites of the drug, but the active metabolites have not been definitively identified. Accordingly, studies were done to determine which metabolites bind to renal mineralocorticoid receptors after SL administration to guinea pigs. The major metabolite found in the steroid receptor fraction of kidney cytosol was 7 alpha-thiomethyl-SL (TM). Incubation of kidney cytosol with varying concentrations (0-100 pmol/l) of aldosterone resulted in the concentration-dependent displacement of TM from the steroid receptor fraction. The steroid receptor fraction from renal nuclei of SL-treated animals contained approximately equal concentrations of TM, 7 alpha-thio-SL (TH), and canrenone (CAN). Incubation of kidney nuclei with aldosterone caused a concentration-dependent displacement of all three metabolites. The results indicate that TM is the major SL metabolite that interacts with cytosolic mineralocorticoid receptors in kidneys, but that TH and CAN may contribute to nuclear receptor binding.

Long-term exposure to norepinephrine results in down-regulation and reduced mRNA expression of pulmonary beta-adrenergic receptors in guinea pigs.

We studied the down-regulation of beta 2-adrenoceptors in guinea pig lung after chronic exposure to a beta-agonist. Guinea pigs were exposed to either norepinephrine (NE) or vehicle for 7 days. The density and affinity of beta-adrenoceptors were determined by Scatchard analysis of [125I]cyanopindolol binding in a lung membrane preparation and beta 1- and beta 2-adrenoceptor subtypes were studied in the presence of 0.1 microM ICI 118,551, a selective beta 2-adrenoceptor antagonist, or 0.1 microM CGP 20712 A, a selective beta 1-adrenoceptor antagonist, respectively. beta 2-Adrenoceptor mRNA was examined by Northern blot analysis. The tissue distribution of beta 2-adrenoceptors and of beta 2-adrenoceptor mRNA in lung after NE infusion were determined using receptor autoradiography and in situ hybridization, respectively. The functional significance of the decrease in beta 2-adrenoceptor was tested by measuring the relaxation response to a beta 2-agonist in isolated parenchymal strips. NE treatment resulted in decreases of 75 +/- 9%, 84 +/- 4%, and 66 +/- 9% of total beta-, beta 1-, and beta 2-adrenoceptors in the lung, compared with control animals. The administration of NE had only minimal effects on the dissociation constant (Kd) of the receptor for the radioligand. beta 2-Adrenoceptors mRNA was decreased by 46 +/- 13% after NE treatment. There was a correspondence between the distribution of beta 2-adrenoceptors and beta 2-adrenoceptor mRNA localization in both control and NE-treated guinea pigs. NE treatment reduced alveolar beta 2-adrenoceptors by 70 +/- 3% and its mRNA expression by 78 +/- 5%.(ABSTRACT TRUNCATED AT 250 WORDS)

Molecular cloning of the guinea-pig histamine H1 receptor gene.

The histamine H1 receptor gene was isolated from a guinea-pig gene library. The gene contains no introns and encodes a protein of 488 amino acid residues. The structure of the guinea-pig histamine H1 receptor is predicted to contain seven putative transmembrane regions, which are similar to those of receptors coupling with GTP binding proteins. Although the third intracellular domain, the predicted binding site for the GTP binding protein, showed only 50% identity with those of the bovine and rat H1 receptors, the expressed guinea-pig H1 receptor was fully able to bind with [3H]mepyramine. Northern blot analysis indicated that the cerebrum, cerebellum, lung, adrenal, intestine, and heart expressed 3.3 kb guinea-pig H1 receptor mRNA. Expression of histamine H1 mRNA of guinea-pig peripheral organs was greater than that of rat organs, suggesting the high sensitivity of guinea-pig organs as to histamine is due to the contents of histamine H1 receptor mRNA. In addition, the lung, adrenal, intestine, and heart expressed 3.9 kb mRNA. In situ hybridization showed that the hippocampus, cerebral cortex, thalamus, and granular layer of the cerebellum each contained a large amount of histamine H1 receptors. Southern blot analysis showed that there was another gene quite similar to the cloned histamine H1 receptor gene.

8‐(3‐Isothiocyanatostyryl)caffeine is a selective, irreversible inhibitor of striatal A2‐Adenosine receptors

8-(3-Isothiocyanatostyryl)caffeine (ISC) was synthesized and shown to inhibit selectively the binding of [(3)H]CGS 21680 (an A(2a)-selective agonist) at adenosine receptors in striatal membranes. The K(i) value at A(2a)-receptors was found to be 110 nM (rat), with selectivity ratios for A(2a) versus A(1)-receptors in rat, guinea pig, bovine, and rabbit striatum of >100-fold. Preincubation of membranes with ISC caused a dose-dependent, irreversible antagonism of the binding of [(3)H]CGS 21680, with an IC(50) value of 3 μM. The irreversibility is likely due to the presence of the chemically reactive isothiocyanate group, since the binding of the corresponding analogue in which the isothiocyanate was replaced with a chloro group was completely reversible. The potency of ISC to irreversibly inhibit the binding of [(3)H]CGS 21680 in several species varied in the order rat ≈ guinea pig > bovine ≈ rabbit. In all four species, binding of the A(1)-selective agonist [(3)H]R-N(6)-phenylisopropyladenosine was not diminished by pre-treatment with 2 μM ISC. The kinetics of irreversible inhibition of rat A(2a)-receptors by 2 μM ISC gave a t(1/2) of approximately 3 min. Following partial inactivation, the remaining rat A(2a)-binding sites retained the same K(d) value as in control membranes for saturation by [(3)H]CGS 21680. Thus, ISC appears to be a selective affinity label for A(2a)- versus A(1)-receptors in the brain.

Tachykinin receptor antagonists inhibit hyperpnea-induced bronchoconstriction in guinea pigs.

We tested the hypothesis that hyperpnea-induced bronchoconstriction (HIB) and hyperpnea-induced bronchovascular hyperpermeability (HIBVH) are mediated through stimulation of NK-1 and NK-2 receptors in guinea pigs. We first established the efficacy and selectivity of (+/-) CP-96,345 (3 mg/kg i.v.) and of SR-48,968 (300 micrograms/kg i.v.) as NK-1 and NK-2 antagonists, respectively. (+/-) CP-96,345 substantially attenuated bronchoconstriction and systemic vascular leak caused by administration of Sar9,Met(O2)11-Substance P (a specific NK-1 agonist), but had no effect upon bronchoconstriction induced by selective NK-2 stimulation with Nle10-Neurokinin A[4-10]. Conversely, SR-48,968 antagonized the bronchoconstrictor response to Nle10-NKA[4-10], right-shifting the dose-response curve by 2 log units, but had no effect on Sar9, Met(O2)11-SP-induced bronchoconstriction. Anesthetized, tracheostomized, opened-chest male Hartley guinea pigs were pretreated with (+/-) CP-96,345 (3 mg/kg i.v.), SR-48,968 (300 micrograms/kg i.v.), or their respective vehicles, and Evans blue dye (30 mg/kg i.v.) to label circulating albumin. 10 min isocapnic dry gas hyperpnea (12 ml/kg, 150 breaths/min) provoked HIB and HIBVH in vehicle-treated animals. (+/-) CP-96,345 reduced the magnitude of HIB by one-half (peak posthyperpnea RL 7.8 +/- 1.9 [SE] times prehyperpnea baseline versus 16.1 +/- 2.6, vehicle-treated; P < or = 0.0001, ANOVA); SR-48,968 blocked HIB more completely (peak posthyperpnea RL 5.1 +/- 1.7 [SE] times prehyperpnea baseline versus 19.3 +/- 2.8, vehicle-treated; P < 0.0001, ANOVA). Neither drug reduced HIBVH. We conclude that dry gas hyperpnea causes bronchoconstriction in guinea pigs through activation of tachykinin receptors. The differential effects of neurokinin receptor blockade on HIB and HIBVH demonstrate that hyperpnea-induced airflow obstruction is not primarily a consequence of hyperpnea-induced bronchovascular leak.

Existence of three subtypes of bradykinin B2 receptors in guinea pig.

We describe the binding of [3H]bradykinin to homogenates of guinea pig brain, lung, and ileum. Analysis of [3H]bradykinin binding kinetics in guinea pig brain, lung, and ileum suggests the existence of two binding sites in each tissue. The finding of two binding sites for [3H]bradykinin in ileum, lung, and brain was further supported by Scatchard analysis of equilibrium binding in each tissue. [3H]Bradykinin binds to a high-affinity site in brain, lung, and ileum (KD = 70-200 pM), which constitutes approximately 20% of the bradykinin binding, and to a second, lower-affinity site (0.63-0.95 nM), which constitutes the remaining 80% of binding. Displacement studies with various bradykinin analogues led us to subdivide the high- and lower-affinity sites in each tissue and to suggest the existence of three subtypes of B2 receptors in the guinea pig, which we classify as B2a, B2b, and B2c. Binding of [3H]bradykinin is largely to a B2b receptor subtype, which constitutes the majority of binding in brain, lung, and ileum and represents the lower-affinity site in our binding studies. Receptor subtype B2c constitutes approximately 20% of binding sites in the brain and lung and is equivalent to the high-affinity site in brain and lung. We suggest that a third subtype of B2 receptor (high-affinity site in ileum), B2a, is found only in the ileum. All three subtypes of B2 receptors display a high affinity for bradykinin, whereas they show different affinities for various bradykinin analogues displaying agonist or antagonist activities.(ABSTRACT TRUNCATED AT 250 WORDS)

Caroverine depresses the activity of cochlear glutamate receptors in guinea pigs: In vivo model for drug-induced neuroprotection?

With the aid of microiontophoretic techniques the action of caroverine, a quinoxaline-derivative, was tested on the receptor-linked depolarisation of the subsynaptic membrane of cochlear afferents. This membrane can be depolarised by the afferent transmitter agonist glutamate, mediated by NMDA and non-NMDA receptors and by acetylcholine, one of the different transmitter substances, released physiologically on axodendritic efferent synapses. Caroverine antagonized the membrane response to glutamate in an enduring but reversible manner. In contrast, the drug exhibited no effect on the depolarising action of acetylcholine. Therefore, the pharmacological profile of caroverine corresponded to the action of selective glutamate receptor antagonists. Since glutamate is likely to be the major mediator of neurotoxicity in the central nervous system, the selective glutamate-antagonism of caroverine is of particular interest, due to its putative neuroprotective competence. Caroverine is currently available clinically in some countries as a spasmolytic drug. Following these results it is proposed to test the drug for clinical efficacy in putatively glutamate-induced, excitotoxic disorders of the brain.

Formula omitted]-alkoxyphenol leukotriene B 4 receptor antagonists: effect of a chroman carboxylic acid.

Several ortho -alkoxyphenols containing a chroman carboxylic acid sidechain have been prepared as antagonists of leukotriene B 4 receptors. These antagonists were compared to their parent alkoxyphenols containing the tetrazole acid sidechain. These chroman containing antagonists retained their binding potency for human neutrophil receptors; however, showed enhanced potency against guinea pig receptors in both in vitro and in vivo systems.

Function of pulmonary M2 muscarinic receptors in antigen-challenged guinea pigs is restored by heparin and poly-L-glutamate.

The effect of heparin and poly-L-glutamate on the function of inhibitory M2 muscarinic autoreceptors on parasympathetic nerves in the lung was tested in antigen-challenged guinea pigs. After antigen challenge, M2 receptor function is decreased, thus increasing release of acetylcholine from the vagus and potentiating vagally induced bronchoconstriction. Guinea pigs were anesthetized, tracheostomized, vagotomized, paralyzed, and ventilated. Electrical stimulation of the vagi caused bronchoconstriction and bradycardia. In controls, pilocarpine attenuated vagally induced bronchoconstriction by stimulating neuronal M2 muscarinic receptors. Conversely, blocking these autoreceptors with gallamine potentiated vagally induced bronchoconstriction. In challenged animals the effects of both drugs were markedly reduced, confirming M2 receptor dysfunction. 20 min after heparin or poly-L-glutamate, the effects of both pilocarpine and gallamine on vagally induced bronchoconstriction were restored, demonstrating recovery of M2 receptor function. Neither heparin nor poly-L-glutamate affected vagally induced responses in control animals. Thus antigen-induced dysfunction of M2 receptors can be reversed by polyanionic polysaccharides (heparin) or polyanionic peptides (poly-L-glutamate). This suggests that a polycationic substance such as eosinophil major basic protein, cationic protein, or peroxidase may be responsible for antigen-induced pulmonary M2 receptor dysfunction.

Characterization of NK-1 receptors in guinea pig and rat brain membranes with NK-1 peptides and a non-peptide antagonist

The major finding of the present investigation is the demonstration of different NK-1 receptors in rat and guinea pig brain membranes with CP 96345 (non-peptide NK-1 antagonist) and R-544 (NK-1 peptide antagonist). We used [ 3H]Sar 9, Me(O 2) 11]SP, the highly selective ligand for NK-1 receptor to compare NK-1 binding sites in rat and guinea pig brain membranes. Scarchard analysis revealed the existence of a single population of [ 3H][Sar 9, Met(O 2) 11]SP binding sites in both preparations. The affinity and the maximal number of bindind sites were found closely similar in rat ( K d 2 nM, B max = 37 fmol/mg protein) and guinea pig brain membranes ( K d = 3 nM, B max = 25 fmol/mg of protein). The order of potency of neurokinins to inhibit [ 3H][Sar 9, Met(O 2) 11]SP binding from rat brain (SP > NKA > NKB) was found different of that observed on guinea pig brain (SP > NKB > NKA). Results obtained with [Sar 9, Met(O 2) 11]SP, [βAla 8]NKA(4–10) and [MePhe 7]NKB suggest that selective agonists cannot discriminate between NK-1 receptors of different species. Using the non-peptide antagonist CP 96345 and the tripeptide R-544, we found that these two NK-1 antagonists discriminate between rat and guinea pig [ 3H][Sar 9, Met(O 2) 11]SP binding sites.

Evaluation of cutaneous algogenic responses to bradykinin and inhibition by bradykinin analogs.

Bradykinin has long been postulated to have a major role in physiologic human pain production. We have developed a method to systematically quantify the algogenic response to bradykinin in human skin by the direct application of bradykinin to suction blister bases. After determination of the pain threshold to bradykinin, we examined the ability of synthetic peptide analogs of bradykinin to block bradykinin-induced pain. One of the six analogs (B4642) inhibited bradykinin-induced pain with at least one concentration in all subjects tested. This result could not be predicted on the basis of inhibition of bradykinin binding to its guinea-pig ileum receptor or to previous studies of bradykinin-induced pain production in animal models. This peptide may have an eventual role as a physiologic pain inhibitor.

Modulation of cardiovascular function by central histamine H 3 receptors in conscious guinea pigs

Modulation of cardiovascular function by central histamine H 3 receptors in conscious guinea pigs

Characterization and tissue distribution of H 3 histamine receptors in guinea pigs by N α-methylhistamine

We have used [ 3H]N α-methylhistamine to characterize H 3-binding in the guinea pig brain and to study its tissue distribution. Kinetic and equilibrium binding experiments indicate a single class of high affinity sites in membranes isolated from guinea pig brain tissue (Kd = 0.4 nM, Bmax = 41 fmol/mg of protein). Competition binding experiments have confirmed that this ligand associates with H 3-receptors and, under the conditions used in these experiments, does not bind to H 1- or H 2-receptors. Although there was some binding in the ileum and large intestine, H 3-binding was found primarily in the central nervous system.

Muscarine receptor types mediating autoinhibition of acetylcholine release and sphincter contraction in the guinea-pig iris

The potencies of several muscarine receptor antagonists in blocking either the autoinhibition of acetylcholine release or the muscarinic contraction of the sphincter muscle upon acetylcholine release were investigated in the guinea-pig iris. The agonist at pre- or postjunctional muscarine receptors was acetylcholine released upon field stimulation (5.5 Hz, 2 min) of the irides preloaded with 14C-choline. The stimulation-evoked 14C-overflow was doubled in the presence of atropine 0.1 mumol/l but unaffected by the agonist (+/-)-methacholine (50 mumol/l). Thus, under the present stimulation conditions, the autoinhibition of acetylcholine release on the guinea-pig iris cholinergic nerves was nearly maximally activated. Isotonic contractions of the irides upon field stimulation consisted of a rapid, atropine (0.1 mumol/l)-sensitive peak phase followed by a sustained contraction which involved a cholinergic and a non-cholinergic stimulation of the sphincter muscle. The M2-selective antagonists methoctramine (10 mumol/l) and gallamine (100 mumol/l) increased both the 14C-overflow and the peak contractions evoked by field stimulation. In contrast, the M3-selective antagonist hexahydrosiladifenidol (0.1-10 mumol/l) failed to affect the evoked 14C-release but concentration-dependently (1-10 mumol/l) reduced the iris contractions. Pirenzepine (10 mumol/l) enhanced the evoked 14C-overflow and inhibited the peak contractions (0.1-10 mumol/l; maximal effect at 10 mumol/l). The low potency of the antagonist at both receptor sites indicates that an M1 muscarine receptor is not involved. The results are consistent with the idea of M2 muscarine receptors mediating autoinhibition of acetylcholine release in the guinea-pig iris and M3-like receptors inducing the contraction of the sphincter muscle.

Binding of dynorphin A and related peptides to κ- and μ-opioid receptors: sensitivity to Na + ions and Gpp(NH)p

We have examined the effects of Na + ions and 5′-guanylyl imidodiphosphate (Gpp(NH)p) on the equilibrium binding of dynorphin A and of a series of related agonist and antogonist peptides to κ- and μ-opioid receptors in guinea pig (κ) and rabbit (μ) cerebellum membrane preparations. The binding to κ sites of dynorphin A and of the peptides displaying agonist properties was strongly inhibited in the presence of 120 mM NaCl and 50 μM Gpp(NH)p. In contrast, a somewhat lower sensitivity to the inhibitory effect of the two allosteric effectors was observed for the analogues of the series showing antagonist properties. The same general behavior, but more marked, was observed at μ sites, at both μ- and κ-opioid receptors. The peptides had biochemical properties (binding sensitivity vs. insensitivity to sodium ions and guanine nucleotides) that correlated well with their biological activities (agonist vs. antagonist) previously determined in in vitro pharmacological bioassays.

Effect of long-term use of reserpine, a sympathetic neuron blocker, on muscarinic cholinergic receptors in the guinea pig nasal mucosa.

To evaluate whether hypofunction of the sympathetic nervous system induced by long-term reserpine treatment affects the parasympathetic system in the nasal mucosa in the receptor level, a receptor binding assay was performed using the nasal mucosa of guinea pigs which had chemical denervation produced by intraperitoneal injection of 0.1 mg/body reserpine once daily for 4 weeks. The maximum number of muscarinic cholinergic receptor bindings to diquinuclidinyl benzilate (3H-QNB) in the nasal mucosa of the reserpine-treated guinea pigs was increased by 46% as compared with that in the control (p less than 0.05). The present experiment shows that hypofunction of the sympathetic system in the nasal mucosa induced by reserpine may affect the parasympathetic system in the nasal mucosa.

Noradrenergic modulation of hypothalamic progestin receptors in female guinea pigs is specific to the ventromedial nucleus

The concentration of estrogen-induced cytosolic progestin receptors (CPRs) in the hypothalamus of α1-noradrenergic antagonist- treated female guinea pigs is reduced relative to non-drug-treated controls. The present study determined where within the hypothalamus this reduction occurs. Ovariectomized, estradiol-treated guinea pigs were given either the α1-noradrenergic antagonist prazosin or vehicle. Microdissected brain regions were assayed for CPR levels. Prazosin caused a selective relative decrease in CPR levels of the ventromedial nucleus of the hypothalamus. No significant effects of prazosin were seen in other hypothalamic or preoptic area nuclei.

Inhibition of sexual behavior in female guinea pigs by a progestin receptor antagonist

The new steroidal progestin receptor antagonist, RU 38486, was used to determine if progesterone-facilitation of sexual behavior in female guinea pigs requires interaction of the hormone with neural progestin receptors. Five milligrams but not 0.5 mg RU 38486 inhibits the expression of sexual behavior in ovariectomized, estrogen-primed guinea pigs treated with 0.1 mg progesterone. This inhibition can be overcome by administration of a large dose of progesterone, suggesting that the drug-effect is specific to the progestin receptor system. RU 38486 binds, in vitro, to progestin receptors and decreases the availability of hypothalamic progestin receptors in estrogen-treated guinea pigs. These studies provide strong evidence that progesterone interaction with intracellular neural progestin receptors mediates the facilitation of sexual behavior by progesterone in female guinea pigs.

Central and peripheral receptors in guinea-pigs exposed to simulated high altitude

Guinea-pigs, chronically exposed to simulated high altitude of 6000 m, showed alterations in some, but not all, neurotransmitter-receptor systems. Thus, peripheral (cardiac) beta-adrenergic receptors were decreased in numbers (Bmax) with an increased affinity although no alterations in endogenous cardiac norepinephrine content were found. In contrast, H1-histamine receptors in the guinea-pig ventricle and brain showed no changes in Bmax or affinity. Dopamine receptors of the caudate nucleus showed no differences between control and altitude-exposed guinea-pigs, although previous reports demonstrated a significant increase (19%) in the content of endogenous dopamine of the caudate. Thus, chronic exposure to simulated high altitude altered beta-adrenergic receptors so that they were more sensitive to small ligand concentrations and less sensitive to large ligand concentrations.