Expression and function of endothelin and its receptors in vascular adventitial fibroblasts

Objective: The adventitia has been recognized to play important roles in vascular oxidative stress, remodelling and contraction. We recently demonstrated that adventitial fibroblasts are able to express endothelin-1 (ET-1) in response to angiotensin II (ANG II). However, the mechanisms by which ANG II induces ET-1 expression are unknown. It is also unclear whether the ET-1 receptors are expressed in the adventitia. We therefore examined the role of oxidative stress in the regulation of ET-1. We also investigated the expression of both the ETA and ETB receptors and the roles of these two types of receptors in collagen synthesis and ET-1 clearance in adventitial fibroblasts. Methods and Results: Adventitial fibroblasts were isolated and cultured from the thoracic mouse aorta. Cells were treated with ANG II (lOOnM), ET-1 (lOpM), NADPH oxidase inhibitor apocynin (lOOfiM), the superoxide anion scavenger tempol (lOOfiM), the ANG II receptor antagonists (100[aM), losartan (AT| receptor) and PD 1233 19 (AT2 receptor), the ET-1 receptor antagonists (lOOuM), BQ123 (ETA receptor) and BQ788 (ETB receptor), and the ETB receptor agonist (lOOnM) Sarafotoxin 6C. ET-1 peptide levels were determined by ELISA, while ETA ,ETB and collagen levels were determined by Western blot. ANG II increased ET-1 peptide levels in a time-dependent manner reaching significance when incubated for 24 hours. NAD(P)H oxidase inhibitor, apocynin, as well as the superoxide scanverger, tempol, significantly reduced ANG Il-induced ET-1 peptide levels while over-expression of SOD1 (endogenous antioxidant enzyme) significantly decreased ANG Il-induced collagen I expression, therefore implicating reactive oxygen species in the mediation of ET-1. ANG II increased ETA receptor protein as well as collagen in a similar fashion, reaching significance after 4, 6, and 24 hours treatment. ANG II induced collagen was reduced while in the presence of the ETA receptor antagonist suggesting the role of the ETa receptor in the regulation of the extracellular matrix. ANG II treatment also increased ETB receptor protein levels in a time-dependent manner. ANG II treatment in the presence of the ETB receptor antagonist significantly increased ET-1 peptide levels. On another hand, the ETB receptor agonist, Sarafotoxin 6C, significantly decreased ET-1 peptide levels. These data implicate the role of the ETb receptor in the clearance of the ET-1 peptide. Conclusion: ANG II-induced increases of ET-1 peptide appears to be mediated by reactive oxygen species derived from NAD(P)H oxidase. Both the ETA and ETB receptors are expressed in adventitial fibroblasts. The ETA receptor subtype mediates collagen I expression, while the ETB receptor may play a protective role through increasing the clearance of the ET- 1 peptide.

Effects of low doses of quinpirole on production of 50 kHz vocalizations in Wistar rats

Rats emit two distinct types of ultrasonic vocalizations in adulthood: 22 kHz (aversive situation), and 50 kHz calls (appetitive situation). The present project is focussed on pharmacological studies of 50 kHz vocalizations. The 50 kHz calls are elicited from dopaminergic activation in the meso limbic pathway and are emitted in such appetitive situations as social contact(s), sexual encounters, food reward, etc. Eighty-five male rats were stereotaxically implanted with bilateral guide cannulae in the nucleus accumbens shell (A= 9.7, L= 1.2, V= 6.7). Quinpirole, a D2/D3 dopaminergic agonist, was injected in low doses to the nucleus accumbens shell in an attempt to elicit 50 kHz vocalizations. A dose response was obtained for the low dose range of quinpirole for six doses: 0.025 Jlg, 0.06 Jlg, 0.12 Jlg, 0.25 Jlg, 0.5 Jlg, and 1.0 Jlg. It was found that only application of the 0.25 Jlg dose of quinpirole and the 7 Jlg dose of amphetamine (positive control) significantly increased the total number of 50 kHz calls (p < 0.006 and p < 0.004 respectively); and particularly significantly increased the frequency modulated type of these calls (p < 0.01, and p < 0.006 respectively). In a double injection procedure, the dose of 0.25 Jlg quinpirole was antagonized with raclopride (D2 antagonist) or U99194A maleate (D3 antagonist) in an attempt to antagonize the response. The 0.25 Jlg dose of quinpirole was successfully antagonized by pre-treatment with an equimolar dose of U99194A maleate (p < 0.008) but not with raclopride. The 7Jlg amphetamine response was also antagonized with an equimolar dose of raclopride. Based on these results, it seems that low doses of quinpirole, particularly the 0.25 Jlg dose, are capable of increasing 50 kHz vocalizations in rats and do so by activation of the D3 dopamine receptor. This is not a biphasic response as seen with locomotor studies. Also noteworthy is the increase in frequency modulated 50 kHz calls elicited by the 0.25 Jlg dose of quinpirole indicating a possible increase in positive affect.

The role of the dopaminergic system in the production of ultrasonic calls in adult rats /

Ultrasonic vocalizations (USV) are emitted by rats in a number of social situations such as aggressive encounters, during sexual behavior, and during play in young rats, situations which are predominantly associated with strong emotional responses. These USV typically involve two distinct types of calls: 22 kHz calls, which are emitted in aversive situations and 50 kHz calls, which are emitted in non-aversive, appetitive situation. The 50 kHz calls are the focus of the present study and to date both the glutamatergic and the dopaminergic systems have been independently implicated in the production of these 50 kHz calls. The present study was conducted to examine a possible relationship between glutamate (GLU) and dopamine (DA) in mediating 50 kHz calls. It was hypothesized that the dopaminergic system plays a mediating role in 50 kHz calls induced by injections ofGLU into the anterior hypothalamic/preoptic area (AHPOA) in adult rats. A total of 68 adult male rats were used in this study. Rats' USV were recorded and analyzed in five experiments that were designed to test the hypothesis: in experiment 1, rats were treated with systemic amphetamine (AMPH) alone; in experiment 2, intra- AHPOA GLU was pretreated with systemic AMPH; in experiment 3, intra-AHPOA GLU was pretreated with intra-AHPOA AMPH; in experiment 4, rats were treated with high and low doses of intra-AHPOA AMPH only; in experiment 5, rats were treated with systemic haloperidol (HAL) as a pretreatment for intra-AHPOA GLU. Analysis of the results indicated that AMPH has a facilitatory effect on 50 kHz USV and that a relationship between DA and GLU in inducing 50 kHz calls does exist. The effect, however, was only observed when DA receptors were antagonized with HAL and was not seen with systemic AMPH pretreatments of intra-AHPOA GLU. The DAGLU relationship at the AHPOA was unclear.

Towards the development of a questionnaire of post transgression motives and behaviours /

The primary purpose of this study was to develop a questionnaire that assesses both forgiveness-seeking motives and behaviours. This questionnaire was based on the premise that, following the commitment of an offense in the context of a relationship, a perpetrator will be motivated to reduce the damage that has taken place. The . ' questionnaire examined several motives that a perpetrator might have for seeking forgiveness. These motives were divided into five proposed domains of posttransgression concerns: God, Self, Victim, Others/Society, and Relationship. Within these domains, the following more specific types of concern were explored: Avoidance of punishment, concern about public image, emotional well-being, self-image, sense of fairness/justice, loss of relationship, loss or gain of power, and loss of ability to trust. The questionnaire also assessed which behaviours (approach and avoidance) a perpetrator might use in order to address these concerns. In addition, this study explored whether or not the severity of the situation and the personality of the perpetrator influenced post transgression motives and behaviours. Participants were 221 individuals from the community and Brock University. They filled out a questionnaire package that assessed personality traits, social desirability, and forgiveness-seeking motives and behaviours. In order to answer items assessing motives and behaviours, participants were asked to imagine themselves as perpetrators in three hypothetical transgression scenarios. These scenarios ranged in severity fi^om low to high. Participants were asked to rate their motives and behaviours both in an immediate time frame (immediately following the transgression) and in the long-term (in order to move on from the situation). Results indicated that the motivation items could be classified into the following subscales:Concern about God, Damaged Self-worth Concerns, Justice Concerns, Impression Management Concerns, Victim and Others Concerns, and Relationship Concerns. The behaviour items formed the following subscales: Approach, Avoidance, Denial and Hiding, and Groveling. Results also indicated differences in motivations and behaviours based on the severity of the situation as well as the personality (assessed using the HEXACO inventory) of the perpetrator.

Formalizing affordances in situation

The representation of a perceptual scene by a computer is usually limited to numbers representing dimensions and colours. The theory of affordances attempted to provide a new way of representing an environment, with respect to a particular agent. The view was introduced as part of an entire field of psychology labeled as 'ecological,' which has since branched into computer science through the field of robotics, and formal methods. This thesis will describe the concept of affordances, review several existing formalizations, and take a brief look at applications to robotics. The formalizations put forth in the last 20 years have no agreed upon structure, only that both the agent and the environment must be taken in relation to one another. Situation theory has also been evolving since its inception in 1983 by Barwise & Perry. The theory provided a formal way to represent any arbitrary piece of information in terms of relations. This thesis will take a toy version of situation theory published in CSLI lecture notes no. 22, and add to the given ontologies. This thesis extends the given ontologies to include specialized affordance types, and individual object types. This allows for the definition of semantic objects called environments, which support a situation and a set of affordances, and niches which refer to a set of actions for an individual. Finally, a possible way for an environment to change into a new environment is suggested via the activation of an affordance.

The role of retinoic acid in long-term memory formation and synaptic plasticity in the mollusc Lymnaea stagnalis

The active metabolite of vitamin A, retinoic acid (RA), is involved in memory formation and hippocampal plasticity in vertebrates. A similar role for retinoid signaling in learning and memory formation has not previously been examined in an invertebrate species. However, the conservation of retinoid signaling between vertebrates and invertebrates is supported by the presence of retinoid signaling machinery in invertebrates. For example, in the mollusc Lymnaea stagnalis the metabolic enzymes and retinoid receptors have been cloned from the CNS. In this study I demonstrated that impairing retinoid signaling in Lymnaea by either inhibiting RALDH activity or using retinoid receptor antagonists, prevented the formation of long-term memory (LTM). However, learning and intermediate-term memory were not affected. An additional finding was that exposure to constant darkness (due to the light-sensitive nature of RA) itself enhanced memory formation. This memory-promoting effect of darkness was sufficient to overcome the inhibitory effects of RALDH inhibition, but not that of a retinoid receptor antagonist, suggesting that environmental light conditions may influence retinoid signaling. Since RA also influences synaptic plasticity underlying hippocampal-dependent memory formation, I also examined whether RA would act in a trophic manner to influence synapse formation and/or synaptic transmission between invertebrate neurons. However, I found no evidence to support an effect of RA on post-tetanic potentiation of a chemical synapse. Retinoic acid did, however, reduce transmission at electrical synapses in a cell-specific manner. Overall, these studies provide the first evidence for a role of RA in the formation of implicit long-term memories in an invertebrate species and suggest that the role of retinoid signaling in memory formation has an ancient origin.