A Mouse Model of Timothy Syndrome: a Complex Autistic Disorder Resulting from a Point Mutation in Cav1.2.

Timothy Syndrome (TS) arises from a point mutation in the human voltage-gated L-type Ca2+ channel (Cav1.2). TS is associated with cardiac arrhythmias and sudden cardiac death, as well as congenital heart disease, impaired cognitive function, and autism spectrum disorders. TS results from a de novo gain-of-function mutation which affects the voltage dependent component of Cav1.2 inactivation. We created a knock-in TS mouse. No homozygous TS mice survived, but heterozygous TS2-NEO mice (with the mutation and the neocassette in situ) had a normal outward appearance and survived to reproductive age. Previously, we have demonstrated that these mice exhibit the triad of Autistic traits. In this paper we document other aspects of these mice including Cav1.2 isoform expression levels, normal physical strength, brain anatomy and a marked propensity towards self-injurious scratching. Gross brain anatomy was not markedly different in TS2-NEO mice compared to control littermates, and no missing structures were noted. The lack of obvious changes in brain structure is consistent with theTS2-NEO mice may provide a significant tool in understanding the role of calcium channel inactivation in both cardiac function and brain development.

Alpha7 neuronal nicotinic receptors as targets for novel therapies to treat multiple domains of schizophrenia.

A number of hypotheses have been put forth to explain the underlying abnormalities of schizophrenia. The widely held dopamine hypothesis suggests that positive symptoms are related to elevated subcortical dopamine transmission and that negative symptoms and cognitive impairments are associated with decreased cortical dopamine function. However, recent evidence suggests broader involvement of serotonergic, glutamatergic and other neurotransmitter systems and a growing body of evidence supports a role for nicotinic cholinergic systems. Based on post-mortem studies, there is a decreased density of neuronal nicotinic receptors (NNRs), especially the alpha7 NNR subtype, in the brains of schizophrenics. The alpha7 NNR subtype is the most abundant in the mammalian brain and has been shown to modulate multiple neuronal pathways that are compromised in schizophrenia, including dopaminergic, serotonergic, glutamatergic and GABAergic pathways. Familial linkage studies have associated regions of chromosome 15, which contains the alpha7 NNR gene, with schizophrenia and polymorphisms have been described in the promoter region of the alpha7 NNR gene. Observations from both animal and human studies that alpha7 NNR agonists can improve positive and negative symptoms as well as cognition to varying degrees further support the involvement of this receptor subtype in multiple deficits of schizophrenia and suggest that it may be feasible to develop novel therapies targeting alpha7 NNRs to treat all domains of the disease.

Serotonergic hyperinnervation and effective serotonin blockade in an FGF receptor developmental model of psychosis.

The role of fibroblast growth factor receptors (FGFR) in normal brain development has been well-documented in transgenic and knock-out mouse models. Changes in FGF and its receptors have also been observed in schizophrenia and related developmental disorders. The current study examines a transgenic th(tk-)/th(tk-) mouse model with FGF receptor signaling disruption targeted to dopamine (DA) neurons, resulting in neurodevelopmental, anatomical, and biochemical alterations similar to those observed in human schizophrenia. We show in th(tk-)/th(tk-) mice that hypoplastic development of DA systems induces serotonergic hyperinnervation of midbrain DA nuclei, demonstrating the co-developmental relationship between DA and 5-HT systems. Behaviorally, th(tk-)/th(tk-) mice displayed impaired sensory gaiting and reduced social interactions correctable by atypical antipsychotics (AAPD) and a specific 5-HT2A antagonist, M100907. The adult onset of neurochemical and behavioral deficits was consistent with the postpubertal time course of psychotic symptoms in schizophrenia and related disorders. The spectrum of abnormalities observed in th(tk-)/th(tk-) mice and the ability of AAPD to correct the behavioral deficits consistent with human psychosis suggests that midbrain 5-HT2A-controlling systems are important loci of therapeutic action. These results may provide further insight into the complex multi-neurotransmitter etiology of neurodevelopmental diseases such autism, bipolar disorder, Asperger's Syndrome and schizophrenia.

Olfactory regulation of the sexual behavior and reproductive physiology of the laboratory mouse: effects and neural mechanisms.

In many species, chemical compounds emitted by conspecifics exert profound effects on reproductive physiology and sexual behavior. This is particularly true in the mouse, where such cues advance and delay puberty, suppress and facilitate estrous cycles, and cause the early termination of pregnancy. They also facilitate sexual behavior and inform mate selection. The mouse has a rich and complex repertoire of social behaviors. The technologies of molecular genetics are well developed in the mouse. Gene expression can be experimentally manipulated in the mouse relatively easily and in a time- and tissue-specific manner. Thus, the mouse is an excellent model in which to investigate the genetic, neural, and hormonal bases by which chemical compounds released by other mice affect physiology and behavior. These chemical cues are detected and processed by the olfactory system and other specialized but less well characterized sensory organs. The sensory information reaches brain regions that regulate hormone levels as well as those that are involved in behavior and alters the function of these brain regions. The effects of these chemical compounds have important implications for the laboratory animal facility as well as for researchers. We begin with an overview of the basic structure and function of the olfactory system and of the connections among brain regions that receive olfactory stimuli. We discuss the effects of chemosensory cues on the behavior and physiology of the organism along with what is known about the neural and hormonal mechanisms underlying these effects. We also describe some of the implications for the laboratory animal facility.

Optimization of laboratory conditions for the study of social behavior.

Social behavior--an action directed toward, or in response to, a member of the same species--is tightly regulated but also highly plastic. It is influenced by many internal (e.g., age, hormonal state, and experience) and external (e.g., time of day, availability of food, encounters with conspecifics) factors. The study of social behavior in the laboratory can be challenging because many facets of social behavior are optimally expressed under specific circumstances. In addition, social behavior is particularly sensitive to environmental factors that are affected by routine animal husbandry. The goal of this article is to review for new investigators and for animal facility staff the major factors that can affect animals' social behavior in the laboratory in order to optimize conditions for the laboratory analysis of social behavior. The authors outline a basic theoretical foundation about the study of social behavior, including the concept of umwelt, an animal's subjective sensory world. They then briefly describe some of the most commonly studied social behaviors and a few examples of the basic methods to analyze these social behaviors. They discuss the potential effects of a facility's husbandry practices on social behavior and how to control these factors as well as possible, with suggestions of several new standard operating procedures toward this end. Although this paper focuses on rodents, the general principles apply to all species. The authors hope that the reader will consider all these factors when designing experiments or working in the animal facility.

The role of the vasopressin 1b receptor in aggression and other social behaviours.

While the importance of vasopressin (Avp) in the neuroendocrine regulation of behaviour is clear, most of Avp's effects on behaviour have been linked to its action via its 1a receptor (Avpr1a) subtype. There is, however, emerging evidence and cross-species consensus that the vasopressin 1b receptor (Avpr1b) is also important in mediating the effects of Avp on behaviour. The Avpr1b is highly expressed in the anterior pituitary where it is thought to play a role in the neuroendocrine response to stress. The Avpr1b is also prominently expressed in the pyramidal cells of the CA2 hippocampal area. Interestingly, in mice, Avpr1b mRNA within the pyramidal neurons of the CA2 field is unaffected by restraint stress or adrenalectomy. Avpr1b knockout mice (--) have provided strong, consistent evidence that the Avpr1b plays a critical role in the regulation of social behaviour. Avpr1b(-/-) mice display reduced levels of social forms of aggression, reduced social motivation and impaired social memory (including the Bruce effect). Avpr1b(-/-) mice, however, have normal main olfactory ability, spatial memory and defensive and predatory behaviours. Mice lacking a functional accessory olfactory system display many of these same behavioural deficits, suggesting that Avpr1b(-/-) mice may have a deficit in the processing, perception and/or integration of olfactory stimuli detected by the accessory olfactory system. We suggest that the role of the Avpr1b is to couple socially relevant accessory olfactory cues with the appropriate behavioural response. Furthermore, given its prominence in the CA2 field of the hippocampus, we hypothesize that Avpr1b may be important for the formation or recall of memories that have an olfactory-based social component.

Inactivation of the oxytocin and the vasopressin (Avp) 1b receptor genes, but not the Avp 1a receptor gene, differentially impairs the Bruce effect in laboratory mice (Mus musculus).

The Bruce effect is a pheromonally mediated process whereby exposure to chemosensory cues from an unfamiliar male terminates pregnancy in a recently mated female. Pharmacological and genetic evidence implicates both oxytocin (Oxt) and vasopressin (Avp) in the regulation of social memory in males, but less work has been done in females. We tested the extent to which the Avp receptors (Avprs) 1a and 1b and Oxt are essential for the Bruce effect, a phenomenon that relies on olfactory memory. Adult female mice were paired with stimulus males and monitored for the presence of sperm plugs. Wild-type, heterozygous, and homozygous knockout (KO) females for either the Avpr1a, Avpr1b, or Oxt genes were randomly assigned to one of the following treatment groups: 1) alone (mate removed, no second exposure to another animal); 2) paired continuously (mate kept with female for 10-14 d); 3) familiar male (mate removed, reintroduced 24 h later); or 4) unfamiliar male (mate removed, BalbC male introduced 24 h later). Regardless of genotype, 90-100% of females in the alone or paired continuously groups became pregnant. The Oxt KO females terminated their pregnancies regardless of whether their original mate or an unfamiliar male was reintroduced. The Avpr1b KO mice failed to terminate pregnancy in the presence of an unfamiliar male. The Avpr1a KO mice exhibited a normal Bruce effect. These data demonstrate that both Oxt and the Avpr1b are critical for the normal expression of the Bruce effect but have different effects on the interpretation of social cues.

The effects of nitric oxide-cGMP pathway stimulation on dopamine in the medial preoptic area and copulation in DHT-treated castrated male rats.

Dopamine (DA) in the medial preoptic area (MPOA) provides important facilitative influence on male rat copulation. We have shown that the nitric oxide-cGMP (NO-cGMP) pathway modulates MPOA DA levels and copulation. We have also shown that systemic estradiol (E(2)) maintains neuronal NO synthase (nNOS) immunoreactivity in the MPOA of castrates, as well as relatively normal DA levels. This effect of E(2) on nNOS probably accounts for at least some of the previously demonstrated behavioral facilitation by intra-MPOA E(2) administration in castrates. Therefore, we hypothesized that stimulation of the MPOA NO-cGMP pathway in dihydrotestosterone (DHT)-treated castrates should restore DA levels and copulatory behaviors. Reverse-dialysis of a NO donor, sodium nitroprusside (SNP), increased extracellular DA in the MPOA of DHT-treated castrates and restored the ability to copulate to ejaculation in half of the animals. A cGMP analog, 8-Br-cGMP, also increased extracellular DA, though not as robustly, but did not restore copulatory ability. The effectiveness of the NO donor in restoring copulation and MPOA DA levels is consistent with our hypothesis. However, the lack of behavioral effects of 8-Br-cGMP, despite its increase in MPOA DA, suggests that NO may have additional mediators in the MPOA in the regulation of copulation. Furthermore, the suboptimal copulation seen in the NO donor-treated animals suggests the importance of extra-MPOA systems in the regulation of copulation.

The acute intoxicating effects of ethanol are not dependent on the vasopressin 1a or 1b receptors.

Studies of the role of vasopressin (Avp) in mediating the effects of ethanol have focused on Avp's role in altering kidney function via its action through the vasopressin 2 receptor. However, alcohol consumption also has central effects that are poorly understood. There is evidence that Avp may mediate ethanol consumption as well as some of ethanol's behavioral effects. Centrally only two Avp receptor subtypes are expressed: the 1a receptor (Avpr1a) and the 1b receptor (Avpr1b). To determine the extent to which these receptors mediate the behavioral effects of alcohol, we used mice with targeted disruptions of either their Avpr1a or Avpr1b gene. We examined the effects of genotype on the acute intoxicating effects of ethanol as well as on voluntary ethanol consumption. Surprisingly, our findings indicate that there is no interaction between either the Avpr1a or Avpr1b and ethanol on motor coordination, hypothermia, mood, or voluntary ethanol consumption.

Social motivation is reduced in vasopressin 1b receptor null mice despite normal performance in an olfactory discrimination task.

In this study, we characterized more thoroughly the social behavior of vasopressin 1b receptor null (V1bR-/-) mice. We confirmed that V1bR-/- males exhibit less social aggression than their wild-type (V1bR+/+) littermates. We tested social preference by giving male subjects a choice between pairs of soiled or clean bedding. In general, V1bR+/+ mice spent significantly more time engaged in chemoinvestigation of these social stimuli than V1bR-/- mice. Male V1bR+/+ mice preferred female-soiled bedding over male-soiled bedding, male-soiled bedding over clean bedding, and female-soiled bedding over clean bedding. In contrast, V1bR-/- males failed to exhibit a preference for any bedding. This difference in behavior is not explained by an anosmic condition as there were no differences between V1bR-/- and V1bR+/+ mice in their abilities to detect a cookie buried in clean bedding, or in their ability to perform in an operant conditioning task using a fully automated liquid dilution olfactometer. In the latter task, male V1bR-/- mice were fully capable of discriminating between male and female mouse urine. The latencies to learn this task did not differ between the two genotypes. Thus, a V1bR-/- male's ability to differentiate between male and female chemosensory cues appears no different than that of a V1bR+/+ male's. We propose that the V1bR plays an important role in social motivation, perhaps by coupling the processing, integration, and/or interpretation of chemosensory cues with the appropriate behavioral response.

Mice lacking D5 dopamine receptors have increased sympathetic tone and are hypertensive.

Dopamine is an important transmitter in the CNS and PNS, critically regulating numerous neuropsychiatric and physiological functions. These actions of dopamine are mediated by five distinct receptor subtypes. Of these receptors, probably the least understood in terms of physiological functions is the D5 receptor subtype. To better understand the role of the D5 dopamine receptor (DAR) in normal physiology and behavior, we have now used gene-targeting technology to create mice that lack this receptor subtype. We find that the D5 receptor-deficient mice are viable and fertile and appear to develop normally. No compensatory alterations in other dopamine receptor subtypes were observed. We find, however, that the mutant mice develop hypertension and exhibit significantly elevated blood pressure (BP) by 3 months of age. This hypertension appears to be caused by increased sympathetic tone, primarily attributable to a CNS defect. Our data further suggest that this defect involves an oxytocin-dependent sensitization of V1 vasopressin and non-NMDA glutamatergic receptor-mediated pathways, potentially within the medulla, leading to increased sympathetic outflow. These results indicate that D5 dopamine receptors modulate neuronal pathways regulating blood pressure responses and may provide new insights into mechanisms for some forms of essential hypertension in humans, a disease that afflicts up to 25% of the aged adult population in industrialized societies.