Cannabinoid exposure during zebra finch sensorimotor vocal learning persistently alters expression of endocannabinoid signaling elements and acute agonist responsiveness

Background  

Previously we have found that cannabinoid treatment of zebra finches during sensorimotor stages of vocal development alters song patterns produced in adulthood. Such persistently altered behavior must be attributable to changes in physiological substrates responsible for song. We are currently working to identify the nature of such physiological changes, and to understand how they contribute to altered vocal learning. One possibility is that developmental agonist exposure results in altered expression of elements of endocannabinoid signaling systems. To test this hypothesis we have studied effects of the potent cannabinoid receptor agonist WIN55212-2 (WIN) on endocannabinoid levels and densities of CB₁ immunostaining in zebra finch brain.     

Role of ionotropic GABA,glutamate and glycine receptors in the tonic and reflex control of cardiac vagal outflow in the rat

Background  

Cardiac vagal preganglionic neurons (CVPN) are responsible for the tonic, reflex and respiratory modulation of heart rate (HR). Although CVPN receive GABAergic and glutamatergic inputs, likely involved in respiratory and reflex modulation of HR respectively, little else is known regarding the functions controlled by ionotropic inputs. Activation of g-protein coupled receptors (GPCR) alters these inputs, but the functional consequence is largely unknown. The present study aimed to delineate how ionotropic GABAergic, glycinergic and glutamatergic inputs contribute to the tonic and reflex control of HR and in particular determine which receptor subtypes were involved. Furthermore, we wished to establish how activation of the 5-HT_1A GPCR affects tonic and reflex control of HR and what ionotropic interactions this might involve.     

Membrane Organization and Dynamics of the G-Protein-Coupled Serotonin<Subscript>1A</Subscript> Receptor Monitored Using Fluorescence-Based Approaches

The G-protein-coupled receptor (GPCR) superfamily represents one of the largest classes of molecules involved in signal transduction across the plasma membrane. Fluorescence-based approaches have provided valuable insights into GPCR functions such as receptor–receptor and receptor–ligand interactions, real-time assessment of signal transduction, receptor dynamics on the plasma membrane, and intracellular trafficking of receptors. This has largely been possible with the use of fluorescent probes such as the green fluorescent protein (GFP) from the jellyfish Aequoria victoria and its variants. We discuss the potential of fluorescence-based approaches in providing novel information on the membrane organization and dynamics of the G-protein-coupled serotonin_1A receptor tagged to the enhanced yellow fluorescent protein (EYFP). These authors contributed equally to the work.     

The <Emphasis Type="Italic">Adhesion</Emphasis> GPCR GPR125 is specifically expressed in the choroid plexus and is upregulated following brain injury

Background  

GPR125 belongs to the family of Adhesion G protein-coupled receptors (GPCRs). A single copy of GPR125 was found in many vertebrate genomes. We also identified a Drosophila sequence, DmCG15744, which shares a common ancestor with the entire Group III of Adhesion GPCRs, and also contains Ig, LRR and HBD domains which were observed in mammalian GPR125.     

Control of synaptic function by endocannabinoid-mediated retrograde signaling

Since the first reports in 2001, great advances have been made towards the understanding of endocannabinoid-mediated synaptic modulation. Electrophysiological studies have revealed that one of the two major endocannabinoids, 2-arachidonoylglycerol (2-AG), is produced from membrane lipids upon postsynaptic Ca²⁺ elevation and/or activation of G_q/11-coupled receptors, and released from postsynaptic neurons. The released 2-AG then acts retrogradely onto presynaptic cannabinoid CB₁ receptors and induces suppression of neurotransmitter release either transiently or persistently. These forms of 2-AG-mediated retrograde synaptic modulation are functional throughout the brain. The other major endocannabinoid, anandamide, mediates a certain form of endocannabinoid-mediated long-term depression (LTD). Anandamide also functions as an agonist for transient receptor potential vanilloid receptor type 1 (TRPV1) and mediates endocannabinoid-independent and TRPV1-dependent forms of LTD. It has also been demonstrated that the endocannabinoid system itself is plastic, which can be either up- or down-regulated by experimental or environmental conditions. In this review, I will make an overview of the mechanisms underlying endocannabinoid-mediated synaptic modulation.     

<Emphasis Type="Italic">N</Emphasis>-Methyl-D-aspartic Acid (NMDA) in the nervous system of the amphioxus <Emphasis Type="Italic">Branchiostoma lanceolatum</Emphasis>

Background  

NMDA (N-methyl-D-aspartic acid) is a widely known agonist for a class of glutamate receptors, the NMDA type. Synthetic NMDA elicits very strong activity for the induction of hypothalamic factors and hypophyseal hormones in mammals. Moreover, endogenous NMDA has been found in rat, where it has a role in the induction of GnRH (Gonadotropin Releasing Hormone) in the hypothalamus, and of LH (Luteinizing Hormone) and PRL (Prolactin) in the pituitary gland.     

Insulin signaling inhibits the 5-HT2C receptor in choroid plexus via MAP kinase

Background

G protein-coupled receptors (GPCRs) interact with heterotrimeric GTP-binding proteins (G proteins) to modulate acute changes in intracellular messenger levels and ion channel activity. In contrast, long-term changes in cellular growth, proliferation and differentiation are often mediated by tyrosine kinase receptors and certain GPCRs by activation of mitogen-activated protein (MAP) kinases. Complex interactions occur between these signaling pathways, but the specific mechanisms of such regulatory events are not well-understood. In particular it is not clear whether GPCRs are modulated by tyrosine kinase receptor-MAP kinase pathways.

Results

Here we describe tyrosine kinase receptor regulation of a GPCR via MAP kinase. Insulin reduced the activity of the 5-HT_2C receptor in choroid plexus cells which was blocked by the MAP kinase kinase (MEK) inhibitor, PD 098059. We demonstrate that the inhibitory effect of insulin and insulin-like growth factor type 1 (IGF-1) on the 5-HT_2C receptor is dependent on tyrosine kinase, RAS and MAP kinase. The effect may be receptor-specific: insulin had no effect on another GPCR that shares the same G protein signaling pathway as the 5-HT_2C receptor. This effect is also direct: activated MAP kinase mimicked the effect of insulin, and removing a putative MAP kinase site from the 5-HT_2C receptor abolished the effect of insulin.

Conclusion

These results show that insulin signaling can inhibit 5-HT_2C receptor activity and suggest that MAP kinase may play a direct role in regulating the function of a specific GPCR.     

Plasticity of histamine H<Subscript>3</Subscript>receptor expression and binding in the vestibular nuclei after labyrinthectomy in rat

Background  

In rat, deafferentation of one labyrinth (unilateral labyrinthectomy) results in a characteristic syndrome of ocular and motor postural disorders (e.g., barrel rotation, circling behavior, and spontaneous nystagmus). Behavioral recovery (e.g., diminished symptoms), encompassing 1 week after unilateral labyrinthectomy, has been termed vestibular compensation. Evidence suggesting that the histamine H₃ receptor plays a key role in vestibular compensation comes from studies indicating that betahistine, a histamine-like drug that acts as both a partial histamine H₁ receptor agonist and an H₃ receptor antagonist, can accelerate the process of vestibular compensation.     

Aromatic interactions impact ligand binding and function at serotonin 5-HT2C G protein-coupled receptors: receptor homology modelling,ligand docking,and molecular dynamics results validated by experimental studies

The serotonin (5-hydroxytryptamine, 5-HT) 5-HT₂ G protein-coupled receptor (GPCR) family consists of types 2A, 2B, and 2C that share ～75% transmembrane (TM) sequence identity. Agonists for 5-HT_2C receptors are under development for psychoses; whereas, at 5-HT_2A receptors, antipsychotic effects are associated with antagonists – in fact, 5-HT_2A agonists can cause hallucinations and 5-HT_2B agonists cause cardiotoxicity. It is known that 5-HT_2A TM6 residues W6.48, F6.51, and F6.52 impact ligand binding and function; however, ligand interactions with these residues at the 5-HT_2C receptor have not been reported. To predict and validate molecular determinants for 5-HT_2C-specific activation, results from receptor homology modelling, ligand docking, and molecular dynamics simulation studies were compared with experimental results for ligand binding and function at wild type and W6.48A, F6.51A, and F6.52A point-mutated 5-HT_2C receptors.     

Activation of muscarinic acetylcholine receptors elicits pigment granule dispersion in retinal pigment epithelium isolated from bluegill

Background

In fish, melanin pigment granules in the retinal pigment epithelium disperse into apical projections as part of the suite of responses the eye makes to bright light conditions. This pigment granule dispersion serves to reduce photobleaching and occurs in response to neurochemicals secreted by the retina. Previous work has shown that acetylcholine may be involved in inducing light-adaptive pigment dispersion. Acetylcholine receptors are of two main types, nicotinic and muscarinic. Muscarinic receptors are in the G-protein coupled receptor superfamily, and five different muscarinic receptors have been molecularly cloned in human. These receptors are coupled to adenylyl cyclase, calcium mobilization and ion channel activation. To determine the receptor pathway involved in eliciting pigment granule migration, we isolated retinal pigment epithelium from bluegill and subjected it to a battery of cholinergic agents.

Results

The general cholinergic agonist carbachol induces pigment granule dispersion in isolated retinal pigment epithelium. Carbachol-induced pigment granule dispersion is blocked by the muscarinic antagonist atropine, by the M₁ antagonist pirenzepine, and by the M₃ antagonist 4-DAMP. Pigment granule dispersion was also induced by the M₁ agonist 4-[N-(4-chlorophenyl) carbamoyloxy]-4-pent-2-ammonium iodide. In contrast the M₂ antagonist AF-DX 116 and the M₄ antagonist tropicamide failed to block carbachol-induced dispersion, and the M₂ agonist arecaidine but-2-ynyl ester tosylate failed to elicit dispersion.

Conclusions

Our results suggest that carbachol-mediated pigment granule dispersion occurs through the activation of M_odd muscarinic receptors, which in other systems couple to phosphoinositide hydrolysis and elevation of intracellular calcium. This conclusion must be corroborated by molecular studies, but suggests Ca²⁺-dependent pathways may be involved in light-adaptive pigment dispersion.

     

Modulatory effect of adenosine receptors on the ascending and descending neural reflex responses of rat ileum

Background

Adenosine is known to act as a neuromodulator by suppressing synaptic transmission in the central and peripheral nervous system. Both the release of adenosine within the small intestine and the presence of adenosine receptors on enteric neurons have been demonstrated. The aim of the present study was to characterize a possible involvement of adenosine receptors in the modulation of the myenteric reflex. The experiments were carried out on ileum segments 10 cm in length incubated in an single chambered organ bath, and the reflex response was initiated by electrical stimulation (ES).

Results

ES caused an ascending contraction and a descending relaxation followed by a contraction. All motility responses to ES were completely blocked by tetrodotoxin, indicating that they are mediated by neural mechanisms. Atropine blocked the contractile effects, whereas the descending relaxation was significantly increased. The A₁ receptor agonist N6-cyclopentyladenosine increased the ascending contraction, whereas the ascending contraction was reduced by the A₁ receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine. Activation of the A₁ receptor further reduced the descending relaxation and the latency of the peristaltic reflex. The A_2B receptor antagonist alloxazine increased ascending contraction, whereas descending relaxation remained unchanged. For A_2A and A₃ receptors, we found contradictory effects of the agonists and antagonists, thus there is no clear physiological role for these receptors at this time.

Conclusions

This study suggests that the myenteric ascending and descending reflex response of the rat small intestine is modulated by release of endogenous adenosine via A₁ receptors.     

Multiple Ligand and Cell-Dependent States of Lateral Mobility of Plasmalemmal G Protein-Coupled Cholecystokinin Receptors

Lateral movement of receptor molecules in the plane of the plasmalemma has important implications for signal transduction and receptor regulation, yet mechanisms affecting such movement are not well understood. We have studied the lateral mobility of the G protein-coupled cholecystokinin (CCK) receptor expressed in the natural milieu of the rat pancreatic acinar cell and in a model cell system, the CHO-CCKR cell, after occupation with fluorescent agonist and antagonist. Lateral diffusion characteristics were distinct in each type of cell and for receptors occupied by each type of ligand, fluorescent agonist, rhodamine-Gly-[(Nle^28,31)CCK-26-33], and fluorescent antagonist, rhodamine-Gly-[(D-Trp³⁰,Nle^28,31)CCK-26-32]-phenethyl ester. Multiple states of mobility were detected for CCK receptors. The slowest population of mobile receptors on the CHO-CCKR cells moved at similar rates when occupied by both antagonist and agonist, while the faster-moving populations moved at a faster rate when occupied with antagonist than with agonist. The fastest component of mobile receptors may reflect unconstrained interactions of the antagonist-occupied receptors with signaling or anchoring structures, while the slowest component may represent the fraction of ligand-occupied receptors that ultimately undergo internalization. The intermediate mobility states may reflect receptor interactions with signal transduction and regulatory machinery. While only a single population of mobile receptors was demonstrable on the acinar cells, increased ligand concentrations (agonist and antagonist) resulted in increased percentages of mobile receptors, suggesting a stoichiometric limitation of immobilizing molecular constraints. Inhibition of protein kinase C had no significant effect on the lateral mobility of agonist-occupied CCK receptors.     

Molecular properties of muscarinic acetylcholine receptors

Muscarinic acetylcholine receptors, which comprise five subtypes (M₁-M₅ receptors), are expressed in both the CNS and PNS (particularly the target organs of parasympathetic neurons). M₁-M₅ receptors are integral membrane proteins with seven transmembrane segments, bind with acetylcholine (ACh) in the extracellular phase, and thereafter interact with and activate GTP-binding regulatory proteins (G proteins) in the intracellular phase: M₁, M₃, and M₅ receptors interact with Gq-type G proteins, and M₂ and M₄ receptors with Gi/Go-type G proteins. Activated G proteins initiate a number of intracellular signal transduction systems. Agonist-bound muscarinic receptors are phosphorylated by G protein-coupled receptor kinases, which initiate their desensitization through uncoupling from G proteins, receptor internalization, and receptor breakdown (down regulation). Recently the crystal structures of M₂ and M₃ receptors were determined and are expected to contribute to the development of drugs targeted to muscarinic receptors. This paper summarizes the molecular properties of muscarinic receptors with reference to the historical background and bias to studies performed in our laboratories.     

Differential inhibition of postnatal brain,spinal cord and body growth by a growth hormone antagonist

Background  

Growth hormone (GH) plays an incompletely understood role in the development of the central nervous system (CNS). In this study, we use transgenic mice expressing a growth hormone antagonist (GHA) to explore the role of GH in regulating postnatal brain, spinal cord and body growth into adulthood. The GHA transgene encodes a protein that inhibits the binding of GH to its receptor, specifically antagonizing the trophic effects of endogenous GH.     

6-S-Alkyl-6-thiohexonolactones and 6-S-Alkyl-6-thiohexitols: Efficient synthesis and new smectic liquid crystal phases

The mesophasic properties of S-alkylthiohexonolactones (d-galactono- and d-mannono-) and the corresponding itol derivatives with general formula Su-SR (R = C₁₁H₂₃) have been investigated. The observation that the synthesized compounds of this series has been studied using crossed polarized optical microscopy. This study involves the discovery of new chiral Smectic A liquid crystal phase through direct comparison with the defective texture of known Smectic A phase of available commercially biphenyls 8CB and 10CB.     

Microglial responses around intrinsic CNS neurons are correlated with axonal regeneration

Background  

Microglia/macrophages and lymphocytes (T-cells) accumulate around motor and primary sensory neurons that are regenerating axons but there is little or no microglial activation or T-cell accumulation around axotomised intrinsic CNS neurons, which do not normally regenerate axons. We aimed to establish whether there was an inflammatory response around the perikarya of CNS neurons that were induced to regenerate axons through a peripheral nerve graft.     

Expression profile of the entire family of <Emphasis Type="Italic">Adhesion</Emphasis>G protein-coupled receptors in mouse and rat

Background  

The Adhesion G protein-coupled receptors (GPCRs) are membrane-bound receptors with long N termini. This family has 33 members in humans. Several Adhesion GPCRs are known to have important physiological functions in CNS development and immune system response mediated by large cell surface ligands. However, the majority of Adhesion GPCRs are still poorly studied orphans with unknown functions.