Smooth muscle does not have a common P2x receptor phenotype: expression, ontogeny and function of P2x1 receptors in mouse ileum, bladder and reproductive systems

The distribution, ontogeny and role of P2x1 receptors were examined in the smooth muscle of the mouse intestine, bladder, and male and female reproductive tracts using P2x1 receptor subtype selective antibodies and contraction studies. P2x1 receptor immunoreactivity showed a heterogeneous distribution in smooth muscle with high levels expressed in adult vas deferens, bladder, arteries and male reproductive organs. In contrast, P2x1, receptors were below the level of detection in the smooth muscle of the ileum and female reproductive tract. P2x1 receptor immunoreactivity was detected at adult levels from birth in the bladder. However, in the vas deferens, immunoreactivity was only detected from 10 days after birth and reached adult levels by approximately 1 month old. A similar pattern of expression was seen in the vesicular seminalis, epididymis, gland of the vas deferens and coagulating gland. Sensitivity to the P2x1 receptor agonist alpha,beta-methylene ATP (alpha,beta-meATP) and P2x1 receptor-deficient mice were used in functional studies to determine the role of P2x1 receptors in the control of smooth muscle. alpha,beta-meATP (100 microM) failed to evoke contractions of the epididymis, or seminal vesicle and P2x1 receptors did not contribute to the control of uterine smooth muscle. In the ileum, alpha,beta-meATP (100 microM) evoked a transient relaxation followed by a contraction. These responses were abolished by the P2 receptor antagonist iso-pyridoxalphosphate-6-azophenyl-2'-5'-disulphonate (iso-PPADS) (30 microM). Relaxant responses were abolished by the adenosine A1 receptor antagonist 1,3-dipropyl-8-cyclopentylxanthine (DPCPX) (1 microM). Contractile responses were reduced by > 80% in the ileum from P2x1 receptor-deficient mice. alpha,beta-meATP-evoked contractions were reduced by approximately 35% by TTX (1 microM) and were unaffected by atropine (10 microM). These studies indicate that P2x1 receptors are not expressed throughout all smooth muscles and that their expression is developmentally regulated. In addition, they provide evidence to suggest that P2x1 receptors are present on pre-synaptic nerve terminals in the enteric nervous system.     

A QTL on rat chromosome 7 modulates prepulse inhibition, a neuro-behavioral trait of ADHD, in a Lewis x SHR intercross

Background  

Attention deficit hyperactivity disorder (ADHD) is a complex neuropsychiatric disorder with a substantial genetic component. The Spontaneously Hypertensive Rats (SHR), considered as a good animal model of ADHD, also show less anxiety-like behaviors than Lewis (LEW) rats. The use of these inbred rat strains led us to the mapping of two quantitative trait loci (QTL), named Ofil1 (on chromosome 4) and Ofil2 (on chromosome 7), related to locomotion in the central and aversive area of an open field. Herein, we examined whether LEW and SHR rats differ in the acoustic startle reflex, a test used to study the neurobiology of anxiety, and in the prepulse inhibition of the startle response, which is known to be impaired in ADHD patients. The effect of the two aforementioned loci on these behavioral responses was also studied.     

Delayed clinical and pathological signs in twitcher (globoid cell leukodystrophy) mice on a C57BL/6 x CAST/Ei background

Modifier genes may account for the phenotypic variability observed in the late-onset forms of globoid cell leukodystrophy (GCL) in humans. In order to begin a search for modifier genes, the effect of genetic background on the clinical and pathological manifestations of GCL was investigated in twitcher mice. Twitcher mice on a C57BL/6 x CAST/Ei background had an increased life span (61.4 +/- 2.5 vs 37.0 +/- 0.6 days), a delayed onset of tremor (24 vs 21 days), and a delayed decline in walking ability compared to C57BL/6 twitcher mice. Pathologically, C57BL/6 x CAST/Ei twitcher mice had fewer lectin-positive globoid cells, less gliosis, and a greater preservation of myelin compared to C57BL/6 twitcher mice under moribund conditions. Similar concentrations of psychosine, the toxic species that accumulates in GCL, were measured by tandem mass spectrometry between moribund C57BL/6 twitcher mice (286.5 pmol/mg protein), 40-day C57BL/6 x CAST/Ei twitcher mice (276.5 pmol/mg), and moribund C57BL/6 x CAST/Ei twitcher mice (247.0 pmol/mg), suggesting that the milder phenotype in CAST/Ei x C57BL/6 twitcher mice did not correlate with less psychosine. In summary, the introduction of modifier genes from the wild, inbred CAST/Ei strain had a phenotypic effect resulting in a significantly slower disease course.     

Bradykinin evoked depolarization of a novel neuroblastoma x DRG neurone hybrid cell line (ND7/23).

Application of bradykinin (Bk) to neuroblastoma x dorsal root ganglion (DRG) neurone hybrid cells (ND7/23) evoked an inward (depolarizing) current associated with an increase in membrane conductance. This response was antagonized by D-Arg0,Hyp3,Thi5,8,D-Phe7-Bk, but was not mimicked by des-Arg9-Bk, indicating the involvement of B2-receptors. The response was unaltered by replacement of extracellular Na+ by N-methylglucamine. Replacement of extracellular Cl by gluconate shifted the estimate reversal potential to a more positive value, while the use of potassium acetate filled recording electrodes shifted the reversal potential to a more negative value, and reduced the response amplitude, indicating the importance of Cl- in the response. This response to Bk was mimicked by the calcium ionophore ionomycin. Bk stimulated the formation of inositol 1,4,5-trisphosphate (IP3), and increased the release of arachidonic acid. In addition, Bk produced an increase in [Ca2+]i, as determined by microspectrofluorimetry. This was due to the release of Ca2+ from intracellular stores, since the response was unaltered when the cells were bathed in Ca(2+)-free solution. In summary, Bk depolarizes ND7/23 cells, probably through the activation of a chloride conductance. It seems likely that this is secondary to the rise in cytosolic Ca2+ concentration, due to the release of Ca2+ from internal stores by IP3. This Ca(2+)-activated chloride response is present in some sensory neurones, although its role in the activation of sensory neurones by Bk is at present unclear.     

Cryptic x; autosome translocation in a boy--delineation of the phenotype

Chromosome X-to-autosome translocations [t(X;A)] are rare rearrangements with an estimated occurrence of 1 to 3 per 10,000 live births. Occurrences of Xq duplications have been observed in male and female subjects in whom the X chromosome segment escapes inactivation and results in functional disomy. We report a case of X;6 translocation in a 7-year-old boy with severe mental retardation, hypotonia, and recurrent respiratory tract infections. High-resolution chromosome analyses (fluorescence in situ hybridization, multiplex ligation probe-dependent amplification, and whole-genome array) revealed a terminal duplication of chromosome X at q28-qter (approximately 3.246 Mb in size) involving gene MECP2 and a terminal deletion (approximately 1.89 Mb) with the breakpoint at 6q27. This is the second report of a boy with a cryptic unbalanced Xq-autosome translocation. This case increases our understanding of mental disability caused by terminal Xq duplication.     

The effect of aluminum on markers for synaptic neurotransmission, cyclic AMP, and neurofilaments in a neuroblastoma x glioma hybridoma (NG108-15)

The effects of the neurotoxin aluminum on markers of synaptic neurotransmission, adenosine 3',5'-monophosphate, and neurofilaments have been evaluated in a neuroblastoma x glioma hybridoma (NG108-15). Cells were exposed for 4 days to 2 mM aluminum lactate, a concentration that did not suppress growth. Compared to controls, the activity of choline acetyltransferase was significantly increased by 37% associated with an up-regulation in enzyme activity (Vmax). Muscarinic receptors, measured by [3H]QNB binding, were reduced by 41%. In contrast, the activities of acetylcholinesterase and glutamate decarboxylase were not significantly changed. Aluminum raised the level of cyclic AMP by 20%, although adenylate cyclase activity was unchanged. Small amounts of both phosphorylated and non-phosphorylated neurofilaments were detected in NG108-15 cells. Aluminum intoxication, however, did not alter the quantity, ultrastructure, or immunoreactivity of neurofilaments. Our results demonstrate the capability of aluminum to produce selected changes in cholinergic markers and levels of cyclic AMP in a rapidly dividing cell line.     

Comparison of a carboxypeptidase E-like enzyme in human, bovine, mouse, Xenopus, shark and Aplysia neural tissue

Several diverse species contain an enzyme with many properties in common with those of bovine carboxypeptidase E (CPE), a neuropeptide processing carboxypeptidase B-like enzyme. This enzyme has been designated EC 3.4.17.10, and is also known as enkephalin convertase and carboxypeptidase H. All tissues that are known to contain bioactive peptides also contain CPE-like enzymatic activity. In Xenopus laevis, enzyme activity is highest in the brain and pituitary, lower in the skin, and undetectable in liver and gut. In Aplysia californica, enzyme activity is highest in the atrial gland, but is also present in moderate amounts in the various neural tissue. CPE extracted from human, bovine, mouse, Xenopus, shark, and Aplysia neural tissue is substantially purified using substrate affinity chromatography and concanavalin A sepharose columns. The partially purified enzyme from all species examined possess very similar enzymatic properties. These properties include a pH optimum of 5.6, a stimulation by cobalt chloride, and an inhibition by chelating agents (1,10-phenanthroline). Arginine-derived active site-directed inhibitors show similar inhibition constants (K_i'ms) towards enzyme from the various species, whereas lysine-derived inhibitors are substantially less potent towards the Aplysia carboxypeptidase than towards enzyme isolated from the other species. The similar properties of the carboxypeptidase isolated from the various species suggests that a CPE-like is involved in the biosynthesis of many peptide neurotransmitters and hormones in a wide range of organisms.     

CYP450, genetics and Parkinson's disease: gene x environment interactions hold the key

The ecogenetic theory contends that most cases of Parkinson's disease (PD) result from the actions of environmental factors in genetically susceptible individuals on a background of normal ageing. This notion is supported by epidemiologic data; family history of PD and exposures to environmental toxins such as pesticides increase risk, while cigarette smoking reduces risk. As a result, polymorphic genes that code for metabolic enzymes have been considered as candidates for conferring differential risk for PD. Given their prominence in xenobiotic metabolism, the cytochrome P450 (CYP) genes have come under great scrutiny. The activity of CYP2D6 is largely determined by genetic variability and common sequence variants exist in human populations that lead to poor metaboliser (PM) phenotypes. These have been extensively studied as genetic risk factors for PD with inconsistent results. However, these studies have disregarded interactive effects (e.g. gene x environment interactions) despite the assertions of the ecogenetic theory. Data from our group and others suggest that the CYP2D6 PM genotype interacts with certain environmental factors such as pesticide exposure and cigarette smoking to confer differential risk for PD. Previous failure to consider such interactions might, in part, explain the inconsistencies observed in the CYP2D6 genetic risk-factor literature. Our data illustrate, using CYP2D6 as an exemplar, that it is crucial to consider both genetic and environmental factors, and their interactions, in any examination of risk factors for PD.     

Expression of a synapse-associated membrane protein, P84/SHPS-1, and its ligand, IAP/CD47, in mouse retina

P84 and integrin associated protein (IAP) are heterophilic binding partners that are expressed in the central nervous system in addition to a variety of other tissues. Both molecules are known to be involved in cell signaling in nonneural tissues. In the retina, both molecules are expressed prominently in plexiform layers, suggesting a possible association with synapses. Here, we examined the cellular expression and ultrastructural localization of the two molecules in the developing mouse retina. Both appeared to be expressed at one or both sides of synaptic sites, although the expression of IAP in the retina precedes that of P84. Examination of transgenic IAP-null retinae revealed a failure of P84 to become associated with synaptic sites, suggesting the interaction of P84 with IAP was necessary for P84's synaptic localization. These findings suggest that the signaling activities of P84 and IAP are localized to sites of synaptic contact in the retina. Thus this pair of synapse-associated molecules represents a bidirectional signaling system that could function to modify synaptic activity or possibly trophic interactions between central neurons.     

Dopamine, learning, and impulsivity: a biological account of attention-deficit/hyperactivity disorder

BACKGROUND: Attention-deficit/hyperactivity disorder (ADHD) affects up to 10% of school-age children. The impulsivity which is seen as its core feature persists over years, yet experimental measures of impulsivity can be altered in a single session. In this study, we tested the theory that both the persistence and the variability of impulsivity could be the result of abnormalities in learning mechanisms and environment. METHOD: We extended an existing model of the role of dopamine in operant conditioning to address the delayed response time task, which is one of the standard tests for impulsivity in ADHD. In this task, subjects choose between immediate responding for a small reinforcer and later responding for a larger one. We studied the influence on impulsivity of four key parameters of the model: The learning rate, discount factor, brittleness, and action bias. RESULTS: The behavior of the model is broadly comparable with electrophysiological (monkey) and behavioral (ADHD and normal) data. Variations in any of the parameters can cause impulsivity. All parameters except the discount factor show inverted U-shaped curves for their effects on impulsivity, suggesting, for example, how either hyper- or hypofunctioning of dopamine can cause impulsivity. The model suggests how decision making can be affected by environmental unpredictability, and thus offers an account of one aspect of the natural history of ADHD. CONCLUSIONS: Some types of ADHD may be caused by specific deficits in reinforcement learning and in the use of learned lessons. Environmental factors can interact with these deficits to delay maturation.     

Effects of 8-OH-DPAT, a 5-HT1A receptor agonist, and DOI, a 5-HT2A/2C agonist, on monosynaptic transmission in spinalized rats

We investigated the effect of the 5-HT_1A receptor agonist (±)-8-hydroxy-2-(di-N-propylamino)tetralin (8-OH-DPAT) and the 5-HT_2A/2C receptor agonist (±)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) on monosynaptic transmission in spinalized rats. 8-OH-DPAT significantly inhibited the excitation of α-motoneurons evoked by monosynaptic transmission without a direct effect on α-motoneuron excitation. DOI potentiated the excitation of α-motoneurons by both direct stimulation and monosynaptic transmission. These results indicate that activation of 5-HT_1A receptors inhibits monosynaptic transmission, whereas activation of 5-HT_2A/2C receptors enhances it.     

Behavioral relationship between autism and fragile x syndrome

Previous researchers have reported autistic features in children with fragile X syndrome. We compared 21 children with pervasive developmental disorders (autism group) to 15 with fragile X syndrome on the Childhood Autism Rating Scale and the Reiss Scales for Children's Dual Diagnosis. The 7 children (47%) with fragile X who scored above the Childhood Autism Rating Scale cut-off (fragile X-autism group) were more impaired than the remaining children (fragile X-no autism) on Childhood Autism Rating Scale subscales related to emotion, visual and listening responses, and communication. The autism group's Reiss scores were higher than fragile X-no autism group, but not fragile X-autism group. Although the Childhood Autism Rating Scale identified almost 50% of children with fragile X as having autism, qualitative differences may exist in specific autistic-like behaviors between children with autism and children with fragile X.     

Zebrafish models for personalized psychiatry: Insights from individual,strain and sex differences,and modeling gene x environment interactions

Currently becoming widely recognized, personalized psychiatry focuses on unique physiological and genetic profiles of patients to best tailor their therapy. However, the role of individual differences, as well as genetic and environmental factors, in human psychiatric disorders remains poorly understood. Animal experimental models are a valuable tool to improve our understanding of disease pathophysiology and its molecular mechanisms. Due to high reproduction capability, fully sequenced genome, easy gene editing, and high genetic and physiological homology with humans, zebrafish (Danio rerio) are emerging as a novel powerful model in biomedicine. Mounting evidence supports zebrafish as a useful model organism in CNS research. Robustly expressed in these fish, individual, strain, and sex differences shape their CNS responses to genetic, environmental, and pharmacological manipulations. Here, we discuss zebrafish as a promising complementary translational tool to further advance patient-centered personalized psychiatry.     

Lovastatin/gemfibrozil myopathy: a clinical, histochemical, and ultrastructural study.

Lovastatin has been used with increasing frequency over the past few years to reduce serum cholesterol. The onset of muscle weakness, one of the most serious side effects of long-term treatment with the drug, constitutes a contraindication to the continuation of therapy and commonly occurs in patients who are also receiving gemfibrozil or cyclosporine. We report the clinical and pathologic findings in a patient treated for hypercholesterolemia with lovastatin and gemfibrozil who developed a rapidly progressive necrotizing myopathy. A 57-year-old woman with hyperlipidemia, treated with lovastatin and gemfibrozil, was admitted to the hospital for evaluation of muscular weakness in her legs and neck. Neurologic examination revealed severe proximal muscle weakness involving both upper and lower extremities as well as proximal muscle tenderness and areflexia in the lower limbs. A biopsy of the quadriceps muscle showed multiple foci of mononuclear cell infiltration with myophagocytosis and slight variation in the size and shape of muscle fibers. Electron microscopy of the affected fibers showed accumulations of subsarcolemmal autophagic lysosomes. The patient's condition dramatically improved after discontinuation of lovastatin-gemfibrozil therapy.     

Host genetic regulation of acute MHV-4 viral encephalomyelitis and acute experimental autoimmune encephalomyelitis in (BALB/cKe x SJL/J) recombinant-inbred mice

In the present report we provide the strain distribution patterns of susceptibility to acute mouse hepatitis virus type-4 (MHV-4) encephalomyelitis, acute experimental allergic encephalomyelitis (EAE) and vasoactive amine sensitivity (VAAS) for 9 (CXJ) recombinant-inbred strains between BALB/cKe (C) and SJL/J(J) mice. We confirm that susceptibility to MHV-4 is not linked to the H-2 complex, and that all strains susceptible to acute EAE have both a responder H-2 haplotype (H-2s or H-2d) and induced (B. pertussis) VAAS. In addition, we provide evidence that susceptibility to acute EAE induction is controlled by an additional presently unmapped locus and that an EAE-like histopathological disease does not usually follow MHV-4 infection intracerebrally in animals susceptible to MHV-4, acute EAE and induced VAAS.     

Depressive-like behavior and stress reactivity are independent traits in a Wistar Kyoto x Fisher 344 cross

Depression is a heritable disorder that is often precipitated by stress. Abnormalities of the stress-reactive hypothalamic-pituitary-adrenal (HPA) axis are also common in depressed patients. In animal models, the forced swim test (FST) is the most frequently used test of depressive-like behavior. We have used a proposed animal model of depression, the Wistar Kyoto (WKY) rat, to investigate the relationship as well as the mode of inheritance of FST behaviors and HPA measures. Through reciprocal breeding of WKY and F344 parent strains and brother-sister breeding of the F1 generation, we obtained 486 F2 animals. Parent, F1 and F2 animals were tested in the FST. Blood samples were collected for determination of basal and stress (10-min restraint) plasma corticosterone (CORT) levels, and adrenal weights were measured. We found that all measures were heritable to some extent and that this heritability was highly sex dependent. Both correlation and factor analyses of the F2 generation data demonstrate that FST behavior and HPA axis measures are not directly related. Thus, the underlying genetic components of depressive-like behavior and HPA axis abnormalities are likely to be disparate in the segregating F2 generation of a WKY x F344 cross.