Effects of imidazolines on neurogenic contraction in isolated urinary bladder detrusor strips from rabbit

Moxonidine and clonidine, which are imidazoline compounds, are sympathetic modulators used as centrally acting antihypertensive drugs. Moxonidine, clonidine, and agmatine produce extensive effects in mammalian tissues via imidazoline recognition sites (or receptors) or α(2)-adrenoceptors. To investigate the effects of imidazolines on the function of the urinary bladder, we tested the effects of moxonidine, clonidine, and agmatine on the neurogenic contraction induced by electric field stimulation, and on the post-synaptic receptors in isolated urinary bladder detrusor strips from rabbit. Both moxonidine at 1.0-10.0?μmol/L and clonidine at 0.1-10.0?μmol/L inhibited electric-field-stimulation-induced contraction in a concentration-dependent manner, but not agmatine (10.0-1000.0?μmol/L). Both moxonidine and clonidine failed to affect carbachol or adenosine-triphosphate-induced contractions; however, 1000.0?μmol/L agmatine significantly increased these contractions. Our study indicates that (i) moxonidine and clonidine produce a concentration-dependent inhibition of the neurogenic contractile responses to electric field stimulation in isolated detrusor strips from male New Zealand rabbits; (ii) post-synaptic muscarinic receptor and purinergic receptor stimulation are not involved in the responses of moxinidine and clonidine in this study; (iii) the inhibitory effects of these agents are probably not mediated by presynaptic imidazoline receptors.     

Insect satiety: sulfakinin localization and the effect of drosulfakinin on protein and carbohydrate ingestion in the blow fly, Phormia regina (Diptera: Calliphoridae)

Sulfakinins, which are satiety factors in invertebrates, have previously been shown to inhibit feeding in the German cockroach and desert locust. This study examines the occurrence of sulfakinin immunoreactivity and the role of sulfakinin as a feeding satiety factor in the black blow fly, Phormia regina. Specifically, this study examines the effect of sulfakinin on two of the blow fly's nutrient requirements (i.e., carbohydrates and proteins). We observed sulfakinin immunoreactive cells in the brains of both male and female flies. We found that drosulfakinin I (DrmSKI, FDDY[SO(3)H]GHMRFa) significantly inhibited carbohydrate feeding by 44% at the most effective dose (10 nmol) in female flies. Statistically, there was no significant effect on males; however, injections of 10 nmol DrmSKI reduced carbohydrate feeding by 34% compared to the sham. Drosulfakinin had no effect on protein feeding and no significant inhibition was detected in females or males. The results of this study lend further support to the idea that carbohydrate and protein feeding are regulated by separate control mechanisms, especially in Calliphoridae.     

Hemodynamic effects of a combination of clonidine and propranolol in conscious cirrhotic rats

The hemodynamic effects of the combination of clonidine and propranolol were studied in conscious rats with portal hypertension owing to secondary biliary cirrhosis. Pressure and blood flow measurements (radioactive microsphere method) were performed in three groups of eight rats before and after drug administration. The combined effects of clonidine (2 micrograms/100 g body wt., i.v.) and propranolol (0.2 mg/min for 10 min) were compared with those observed after administration of either clonidine alone or propranolol alone. The association of clonidine and propranolol induced significant decreases in portal pressure (30%) and portal tributary blood flow (43%), the magnitude of these changes being significantly more marked than that after administration of either clonidine alone (12 and 20%, respectively) or propranolol alone (16 and 17%, respectively). After the combination, no significant change in arterial pressure was observed, but cardiac output significantly decreased and systemic vascular resistance significantly increased. Renal blood flow decreased to a similar extent (40%) in the three groups. These findings indicate that the combination of clonidine and propranolol is more effective for reversing splanchnic hemodynamic changes than clonidine alone or propranolol alone. The additive effects of this association are in agreement with the action of clonidine and propranolol at different levels (central and peripheral) and on different receptors (alpha and beta). It suggests that an increase in sympathetic activity may play a major role in hemodynamic changes observed in experimental cirrhosis.     

G proteins control diverse pathways of transmembrane signaling

Hormones, neurotransmitters, and autacoids interact with specific receptors and thereby trigger a series of molecular events that ultimately produce their biological effects. These receptors, localized in the plasma membrane, carry binding sites for ligands as diverse as peptides (e.g., glucagon, neuropeptides), lipids (e.g., prostaglandins), nucleosides and nucleotides (e.g., adenosine), and amines (e.g., catecholamines, serotonin). These receptors do not interest directly with their respective downstream effector (i.e., an ion channel and/or an enzyme that synthesizes a second messenger); rather, they control one or several target systems via the activation of an intermediary guanine nucleotide-binding regulatory protein or G protein. G proteins serve as signal transducers, linking extracellularly oriented receptors to membrane-bound effectors. Traffic in these pathways is regulated by a GTP (on)-GDP (off) switch, which is regulated by the receptor. The combination of classical biochemistry and recombinant DNA technology has resulted in the discovery of many members of the G protein family. These approaches, complemented in particular by electrophysiological experiments, have also identified several effectors that are regulated by G proteins. We can safely assume that current lists of G proteins and the functions that they control are incomplete.     

Multitrophic interactions of the silverleaf whitefly,host plants,competing herbivores,and phytopathogens

Our laboratory found that silverleaf whitefly (SLW; Bemisia argentifolii Bellows & Perring) feeding alters host plant physiology and chemistry. The SLW induces a number of host plant defenses, including pathogenesis-related (PR) protein accumulation (e.g., chitinases, beta-1,3-glucanases, peroxidases, chitosanases, etc.). Induction of the PR proteins by SLW feeding occurs in various plant species and varieties. The extent and type of induction is dependent on a number of factors that include host plant growing conditions, the length of time the host plant is exposed to SLW feeding, the plant variety, and SLW population densities. The appearance of PR proteins correlates well with reduced infestations of conspecific insect herbivore competitors. Greenhouse and field experiments in which herbivore competitors (cabbage looper, Trichoplusia ni; leaf miner, Liromyza trifolii) were placed on plants previously exposed to SLW feeding demonstrated behavioral differences (oviposition, feeding preferences) and reduced survival rates and development times of these insects. The interaction was asymmetrical, i.e., SLW infestations of plants previously exposed to leaf miners had little or no effect on SLW behavior (oviposition). Induction of plant-defensive proteins by SLW feeding was both local (at the feeding site) and systemic (uninfested leaves distant to the feeding site). There are interactions between diseases such as tomato mottle virus (ToMoV; a geminivirus) and the host plant and SLW. PR proteins were induced in tomato plants infected with ToMoV much as they were via non-viruliferous SLW feeding. The presence of ToMoV in tomato plants significantly increased the number of eggs produced by SLW females. Experiments using tomato plants, powdery mildew (PM), and tobacco mosaic virus (TMV) show that whitefly infestations can affect plant pathogen relationships but the effects vary among pathogen types. Enzyme analyses prior to pathogen inoculation showed that whitefly treatment significantly increased the activities of foliar chitinase and peroxidase. Evaluation of pathogen growth 3 weeks after inoculation showed that whitefly feeding significantly reduced the incidence of PM. However, TMV levels evaluated by ELISA were not significantly affected by whitefly feeding. Six weeks after inoculation with pathogens, the chitinase and peroxidase activities were still elevated in plants initially fed on by whiteflies but continuing pathogen infection had no effect on these enzymes. The possibility that geminivirus infection and/or SLW infestations isolate the host plant for the selected reproduction of the virus and the insect is discussed. Multitrophic cascade effects may contribute to the successful eruptive appearance of SLW on various crops, ranking them as a major pest. They may explain the general observation that when SLW infest a host plant there are few if any competing insect herbivores and pathogens found in the host. However, the results indicate that certain SLW-virus relationships could be mutualistic.     

α2-Adrenergic Receptors in Neuronal and Glial Cultures: Characterization and Comparison

Membranes prepared from either neuronal or glial cultures contain alpha 2-adrenergic receptors as determined by the characteristics of [3H]yohimbine [( 3H]YOH) binding. The binding was rapid, reversible, saturable, dependent on the protein concentration used, and reached equilibrium by 5 min in membranes from both neuronal and glial cultures. Scatchard analyses of saturation isotherms revealed similar KD values of 13.7 +/- 1.35 nM (n = 10) for neuronal cultures and 18.42 +/- 2.34 nM (n = 10) for glial cultures. Glial cultures contained many more binding sites for [3H]YOH than neuronal cultures, having a Bmax of 1.6 +/- 0.33 pmol/mg protein (n = 10) compared with 0.143 +/- 0.018 pmol/mg protein (n = 10) in neurons. Drugs selective for alpha 2-adrenergic receptors were the most effective displacers of [3H]YOH binding in both neuronal and glial cultures, i.e., the alpha 2-adrenergic antagonists rauwolscine and yohimbine were better displacers than the other catecholamine antagonists prazosin, corynanthine, or propranolol. The agonists showed the same pattern with the alpha 2-selective drugs clonidine and naphazoline being the most effective competitors for the [3H]YOH site. GTP and its nonhydrolyzable analog. 5'-guanylyl-imidodiphosphate, were able to lower the affinity of the alpha 2-receptors for agonists but not antagonists in membranes from both neuronal and glial cultures, suggesting that the receptors are linked to a G protein in both cell types. The presence of alpha 2-adrenergic receptors in neuronal cultures was also substantiated by light microscopic autoradiography of [3H]YOH binding. In summary, we have demonstrated that both neuronal and glial cultures contain alpha 2-adrenoceptors.     

Neurotrophins, airway smooth muscle and the fetal breathing-like movements

Central nervous system and skeletal muscles secrete a group of polypeptide hormones called neurotrophins (NTs). More recent studies show that NTs and their receptors are also expressed in the lung, suggesting a role for NTs in lung development. To examine the role of NTs during normal and diseased lung organogenesis, we employed wild-type and amyogenic mouse embryos (designated as Myf5-/-:MyoD-/-). Amyogenic embryos completely lacked skeletal muscles and were not viable after birth due to the respiratory failure secondary to lung hypoplasia. To examine the importance of lung-secreted NTs during normal and hypoplastic lung organogenesis, immunohistochemistry was employed. Distribution of NTs and their receptors was indistinguishable between normal and hypoplastic lungs. To further examine the importance of non-lung-secreted NTs (e.g., from the skeletal muscle and CNS) in lung organogenesis, in utero injections of two NTs were performed. The exogenously introduced NTs (i.e., non-lung-secreted) did not appear to improve development of the lung in amyogenic embryos. Moreover, immunohistochemistry showed significantly reduced number of airway smooth muscle cells (ASMCs) in hypoplastic lungs of amyogenic embryos, suggesting that the number of ASMCs is primarily regulated by the fetal breathing-like movements (i.e., mechanical factors).     

Behavioral correlates of supplementary feeding of wildlife: Can general conclusions be drawn?

Supplementary feeding is a common, but controversial, tool in wildlife management, because it can benefit both humans and wildlife (e.g., increased wildlife densities), but has certain downsides (e.g., increased disease transmission). For species that are often involved in human-wildlife conflicts, two opposing paradigms with respect to supplementary feeding exist, i.e., (i) that supplementary feeding is efficient to lure animals away from undesired places (i.e., diversionary feeding; hypothesis 1), and (ii) that supplementary feeding stimulates ‘nuisance’ behavior (i.e., increased tolerance for humans and selection for human facilities; hypothesis 2). We formulated an alternative hypothesis (hypothesis 3); i.e., that behavioral variation among individuals dilutes population-wide, general patterns with respect to supplementary feeding. Based on GPS relocation data and resource selection functions, we show that neither of the two opposing management paradigms (hypothesis 1 and 2) hold in a particularly ‘conflict rich’ species, the brown bear (Ursus arctos), because individual variation in selection behavior with respect to supplementary feeding diluted population-wide patterns (hypothesis 3), even under very different environmental contexts (Sweden vs. Slovenia; i.e., different human and bear population density, history and intensity of supplementary feeding, topography, etc.). Our results emphasize that individual variation is an important component of behavioral ecology and should be considered in wildlife management and conservation.     

Intracerebroventricular injection of antibodies directed against Gs alpha enhances the supraspinal antinociception induced by morphine, beta-endorphin and clonidine in mice.

The intracerebroventricular (i.c.v.) injection of antisera directed against different sequences of Gs alpha to mice enhanced the antinociceptive potency of the opioids morphine, beta h-endorphin-(1-31) and of the alpha 2-agonist clonidine when studied 24 h later in the tail-flick test. The activity of DAGO, DADLE, DPDPE and [D-Ala2]-Deltorphin II remained unchanged after that treatment. Cholera toxin (0.5 microgram/mouse, i.c.v.), agent that impairs the receptor regulation of Gs transducer proteins promoted comparable changes in the supraspinal analgesia induced by these substances. Six days after a single i.c.v. injection (0.5 microgram/mouse) of pertussis toxin the antinociceptive activity of all the opioids and clonidine appeared diminished. It is concluded that opioids and clonidine promote analgesia after binding to receptors functionally coupled to Gi/G(o) proteins, moreover, the activity of morphine, beta-endorphin and clonidine in this test seems to be counteracted by a process involving activation of Gs alpha transducer proteins.     

Inhibition of lipolysis in hamster adipocytes with selective alpha-adrenergic stimuli. Functional characterization of the alpha-receptor

This communication shows the relative potencies of the alpha-agonists clonidine, methoxamine, methyl norepinephrine and phenylephrine in producing inhibition of lipolysis. At cell densities greater than 15 mg cell/ml lipolysis activated by either 1-methyl-3-isobutyl xanthine or adenosine deaminase was inhibited by alpha-adrenergic stimuli with a rank order of potency of clonidine greater than methoxamine greater than methyl norepinephrine; phenylephrine produced a further stimulation of lipolysis. At the same cell density isoproterenol-accelerated lipolysis was inhibited by alpha-adrenergic stimuli with a rank order of potency of phenylephrine greater than methoxamine greater than clonidine greater than methyl norepinephrine. When the density of fat cells was reduced to less than 5 mg/ml, clonidine was a more effective inhibitor of isoproterenol-activated lipolysis thatn phenylephrine. Lipolysis that was activated by dibutyryl cyclic AMP, ACTH or cholera enterotoxin was not reduced by any alpha-adrenergic agent. Under conditions when clonidine failed to inhibit catecholamine-activated lipolysis (i.e., at cell densities greater than 15 mg/ml), it failed to antagonize the antilipolytic activity of phenylephrine. The antilipolytic activities of clonidine and phenylephrine were most effectively antagonized by the blocking drugs phentolamine and yohimbine; in contrast, phenoxybenzamine and prazosin were less effective blockers. These data indicate that the alpha-adrenergic receptor on hamster fat cells is similar to presynaptic alpha-adrenergic receptors. The data further suggest the possibility that phenylephrine may exert its action through a separate alpha-adrenergic receptor mechanism.     

Does conditioned taste aversion learning in the pond snail Lymnaea stagnalis produce conditioned fear?

In conditioned taste aversion (CTA) training performed on the pond snail Lymnaea stagnalis, a stimulus (the conditional stimulus, CS; e.g., sucrose) that elicits a feeding response is paired with an aversive stimulus (the unconditional stimulus, US) that elicits the whole-body withdrawal response and inhibits feeding. After CTA training and memory formation, the CS no longer elicits feeding. We hypothesize that one reason for this result is that after CTA training the CS now elicits a fear response. Consistent with this hypothesis, we predict the CS will cause (1) the heart to skip a beat and (2) a significant change in the heart rate. Such changes are seen in mammalian preparations exposed to fearful stimuli. We found that in snails exhibiting long-term memory for one-trial CTA (i.e., good learners) the CS significantly increased the probability of a skipped heartbeat, but did not significantly change the heart rate. The probability of a skipped heartbeat was unaltered in control snails given backward conditioning (US followed by CS) or in snails that did not acquire associative learning (i.e., poor learners) after the one-trial CTA training. These results suggest that as a consequence of acquiring CTA, the CS evokes conditioned fear in the conditioned snails, as evidenced by a change in the nervous system control of cardiac activity.     

Both ligand- and cell-specific parameters control ligand agonism in a kinetic model of g protein-coupled receptor signaling

G protein-coupled receptors (GPCRs) exist in multiple dynamic states (e.g., ligand-bound, inactive, G protein-coupled) that influence G protein activation and ultimately response generation. In quantitative models of GPCR signaling that incorporate these varied states, parameter values are often uncharacterized or varied over large ranges, making identification of important parameters and signaling outcomes difficult to intuit. Here we identify the ligand- and cell-specific parameters that are important determinants of cell-response behavior in a dynamic model of GPCR signaling using parameter variation and sensitivity analysis. The character of response (i.e., positive/neutral/inverse agonism) is, not surprisingly, significantly influenced by a ligand's ability to bias the receptor into an active conformation. We also find that several cell-specific parameters, including the ratio of active to inactive receptor species, the rate constant for G protein activation, and expression levels of receptors and G proteins also dramatically influence agonism. Expressing either receptor or G protein in numbers several fold above or below endogenous levels may result in system behavior inconsistent with that measured in endogenous systems. Finally, small variations in cell-specific parameters identified by sensitivity analysis as significant determinants of response behavior are found to change ligand-induced responses from positive to negative, a phenomenon termed protean agonism. Our findings offer an explanation for protean agonism reported in beta2--adrenergic and alpha2A-adrenergic receptor systems.     

Isolation of human cytomegalovirus intranuclear capsids, characterization of their protein constituents, and demonstration that the B-capsid assembly protein is also abundant in noninfectious enveloped particles.

Two types of intranuclear capsids have been recovered from human cytomegalovirus (HCMV, strain AD169)-infected cells. By analogy with strain Colburn (simian CMV) particles, these have been designated as A- and B-capsids. Both types of capsids are composed of proteins with molecular weights of 153,000 (major capsid protein), 34,000 (minor capsid protein), 28,000, and 11,000 (smallest capsid protein). In addition to these species, B-capsids contain a 36,000-molecular-weight (36K) protein which has been designated as the HCMV "assembly protein," based on its similarities to counterparts in strain Colburn CMV (i.e., 37K protein) and herpes simplex virus (i.e., VP22a/p40/NC-3/ICP35e). Peptide comparisons established that the assembly protein of HCMV B-capsids and the 36K protein that distinguishes HCMV noninfectious enveloped particles from virions are the same, providing direct evidence that noninfectious enveloped particles are enveloped B-capsids.     

Contribution of AMPA/kainate receptors in the rostral ventrolateral medulla to the hypotensive and sympathoinhibitory effects of clonidine

The depressor and sympathoinhibitory effect of the imidazoline drug clonidine is reported to be associated with functional states of the central glutamate receptors. The rostral ventrolateral medulla (RVLM) has been recognized as a specific target area for mediating the central depressor mechanism of clonidine. The objective of this study was to determine the role of the glutamate receptor subtype alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/kainate receptor within the RVLM in clonidine-induced depressor and sympathoinhibitory action in anesthetized normotensive rats. Unilateral microinjection of 200 pmol of 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), a potent AMPA/kainate receptor antagonist, into the RVLM completely abolished the pressor action evoked by AMPA (5 pmol) without affecting the pressor action of N-methyl-D-aspartate (20 pmol). Pretreatment with intra-RVLM injection of CNQX (20 and 200 pmol) dose dependently attenuated the reduction in blood pressure (BP), heart rate (HR), and renal sympathetic nerve activity (RSNA) elicited by intra-RVLM clonidine (5 nmol) or intravenous clonidine (10 microg/kg), while 2 pmol of CNQX did not alter clonidine-induced cardiovascular action. Furthermore, the decreases in BP, HR, and RSNA evoked by intravenous clonidine (10 microg/kg) or intra-RVLM clonidine (5 nmol) were reversed when CNQX (20 and 200 pmol) was subsequently injected into the RVLM. In conclusion, these data show that blockade of AMPA/kainate receptors in the RVLM significantly antagonizes decreases in BP, HR, and sympathetic activity induced by clonidine, suggesting that the AMPA/kainate receptors within the RVLM contribute to the depressor and sympathoinhibitory effect of clonidine.     

A Comparative Test of Adaptive Explanations for Hypsodonty in Ungulates and Rodents

Hypsodonty has long been recognized as an adaptation for grazing: grazing is suggested to increase tooth wear due to endogenous (e.g., fiber, silica) and/or exogenous (e.g., dust, grit) properties of ingested food. However, it is unknown whether tooth crown height is correlated with the mastication of high fiber or silica in grasses, the ingestion of external abrasives, or both. Furthermore, comparative studies of hypsodonty have not explicitly taken into account phylogenetic biases due to shared ancestry in tooth morphology and/or feeding behavior. This study highlights the relationship between molar crown height and feeding habits in African ungulates and South American rodents when phylogenetic effects are controlled. Among ungulates, high hypsodonty indices are significantly associated with specific plant and foraging height preferences, while habitat and climate show no correlation with tooth crown height. For rodents, grass-eating species are significantly more hypsodont than frugivorous or folivorous species, and arboreal rodents are less hypsodont than terrestrial species. These results as well as those of a posteriori analyses controlling for aspects of the behavioral ecology (e.g., grass-eating, substrate preference) of the sample species confirm the role of both diet and grit in shaping the evolution of cheek tooth crown height in herbivorous mammals.     

Both Ligand- and Cell-Specific Parameters Control Ligand Agonism in a Kinetic Model of G Protein–Coupled Receptor Signaling

G protein–coupled receptors (GPCRs) exist in multiple dynamic states (e.g., ligand-bound, inactive, G protein–coupled) that influence G protein activation and ultimately response generation. In quantitative models of GPCR signaling that incorporate these varied states, parameter values are often uncharacterized or varied over large ranges, making identification of important parameters and signaling outcomes difficult to intuit. Here we identify the ligand- and cell-specific parameters that are important determinants of cell-response behavior in a dynamic model of GPCR signaling using parameter variation and sensitivity analysis. The character of response (i.e., positive/neutral/inverse agonism) is, not surprisingly, significantly influenced by a ligand's ability to bias the receptor into an active conformation. We also find that several cell-specific parameters, including the ratio of active to inactive receptor species, the rate constant for G protein activation, and expression levels of receptors and G proteins also dramatically influence agonism. Expressing either receptor or G protein in numbers several fold above or below endogenous levels may result in system behavior inconsistent with that measured in endogenous systems. Finally, small variations in cell-specific parameters identified by sensitivity analysis as significant determinants of response behavior are found to change ligand-induced responses from positive to negative, a phenomenon termed protean agonism. Our findings offer an explanation for protean agonism reported in β₂--adrenergic and α_2A-adrenergic receptor systems.     

Interferon action: effects of mouse alpha and beta interferons on rosette formation, phagocytosis, and surface-antigen expression of cells of the macrophage-type line RAW 309Cr.1

Treatment with an essentially pure mouse α or β interferon boosts the binding and phagocytosis of opsonized sheep red blood cells by cells of the murine macrophage-like cell line RAW 309Cr.l. The kinetics and the dose dependence of the effects of the two interferons are very similar. The effects depend on continued RNA and protein synthesis, and they diminish after the removal of interferon from the medium. Studies with agents specifically binding FcRI receptors (i.e., IgG2a) and FcRII receptors (i.e., the Fab fragment of the antireceptor monoclonal antibody 2.4G2) revealed a three- to fivefold increase in the level of FcRI receptors per cell and an about twofold increase in that of FcRII receptors per cell after treatment with interferon. The enhanced binding and phagocytosis of opsonized sheep red blood cells by interferon treatment are apparently a consequence of the increased number of Fc receptors. As revealed by studies involving the binding to the cells of labeled monoclonal antibodies to several cell surface antigens, the level of the H-2D^d surface antigen is also selectively increased three- to fourfold in the cells after exposure to interferon.     

Cholera toxin and pertussis toxin on opioid- and alpha 2-mediated supraspinal analgesia in mice.

Cholera toxin, an agent that impairs the function of Gs transducer proteins, was injected (0.5 microgram/mouse, icv) and the antinociceptive activity of opioids and clonidine was studied 24h later in the tail-flick test. In these animals, an enhancement of the analgesic potency of morphine, beta-endorphin and clonidine could be observed. Cholera toxin did not modify the antinociception evoked by the enkephalin derivatives DAGO and DADLE. Pertussis toxin that catalyses the ADP ribosylation of alpha subunits of Gi/Go regulatory proteins was given icv (0.5 microgram/mouse). This treatment reduced the analgesic effect of opioids and clonidine. However, while the analgesia elicited by DAGO, DADLE and clonidine was greatly decreased, the effect of morphine and beta-endorphin was reduced to a moderate extent. It is concluded that Gi/Go regulatory proteins functionally coupled to opioid and alpha 2 receptors are implicated in the efficacy displayed by opioids and clonidine to produce supraspinal analgesia. Moreover, these two receptors are susceptible to regulation by a process that might involve a Gs protein.