Alan Sved - US grants

The central nervous system plays a key role in the regulation of the cardiovascular system and altered central neural control of blood pressure (BP) may participate in the initiation or maintenance of hypertension. Three brainstem areas are of key importance in BP regulation: the rostral ventrolateral medulla (RVL), the caudal ventrolateral medulla (CVL), and the nucleus tractus solitarius (NTS). Recent data from pharmacological studies indicate that tonically active GABAergic synapses in each of these regions influence BP. The goal of the experiments described in this proposal is to study the role of GABAergic neurotransmission in the NTS, RVL, and CVL in the regulation of BP and to determine whether altered GABAergic neurotransmission in these regions may be involved in the pathogenesis of hypertension. The first set to experiments will utilize a pharmacological approach to study how drugs which influence GABAergic neurotransmission affect blood pressure and heart rate when administered directly into the NTS, RVL, or CVL. These studies will focus primarily on the effects of indirect-acting GABA agonists, since the responses to such drugs should reflect the ongoing level of GABAergic neurotransmission. Responses to these drugs will be compared in hypertensive and normotensive rats. The second set of studies will use a neurochemical approach to examine similar questions. In these studies, the rate of GABA synthesis (which presumably reflects the rate of GABA released into the synapse) will be determined by measuring the accumulation of GABA following local inhibition of its metabolism. A push-pull perfusion technique will be used to confirm that observed changes in synthesis do reflect changes in GABA release. Using these techniques, it will be determined whether changes in cardiovascular afferent activity affect GABAergic neurotransmission in the NTS, RVL, or CVL. These techniques will also be applied to determine whether GABAergic neurotransmission is altered in the NTS, RVL, or CVL in experimental hypertension in rats. These studies will provide a comprehensive analysis of GABAergic neurotransmission in the NTS, RVL, and CVL as related to the regulation of blood pressure and the pathogenesis of hypertension. It is expected that these studies will provide new and important information on the role of the brain in hypertension, and may therefore offer new insights into the treatment or prevention of hypertension.

The regulation of blood pressure is a complex process, involving the integrated action of many systems. The central nervous system is a key component of this regulatory process, and one site in the brain which has received considerable attention in this regard is the nucleus tractus solitarius (NTS). This nucleus is the site of termination of the vast majority of baroreceptor afferent inputs, and is therefore the first site in the central nervous system involved in the mediation and integration of the baroreceptor reflex. Bilateral destruction of the NTS in rats leads rapidly to severe hypertension, resulting in death within several hours. The hypertension is caused by increased sympathoadrenal activity and increased release of vasopressin into the circulation. In contrast, it has recently been observed that if rats are pretreated to prevent the acute phase of this hypertension, they survive and chronically maintain a normal resting blood pressure, although baroreceptor input and integration should be totally abolished. The present proposal is designed to examine the mechanisms by which normal resting blood pressure is restored in rats with bilateral lesions of the NTS. The first group of experiments is designed to verify that baroreceptor reflexes are abolished in rats with chronic lesions of the NTS. Following this, experiments will examine whether sympathetic activity and vasopressin release are chronically enhanced in rats with NTS lesions. Experiments will then address (a) how sympathetic activity and vasopressin release return to normal following NTS lesions or (b) how normal blood pressure is restored in the presence of chronically enhanced sympathetic outflow and vasopressin release. Thus, these studies will examine the ability of the cardiovascular system to adapt to disruption of the central neural control of blood pressure caused by destruction of the NTS. It is expected that these studies will provide important new information on the regulation of blood pressure exerted by the brain, and the integration of the many systems normally involved in the regulation of blood pressure.

The central nervous system (CNS) plays a key role in regulation of arterial blood pressure and an abnormality of the central neural control may result in hypertension or predispose an individual to develop hypertension in response to other factors. Studies conducted the current grant period have highlighted the role of the CNS in baroreceptor- independent long-term control of blood pressure. These studies have focused on the role of neurons in the rostral ventrolateral medulla (RVLM) in maintaining the tonic drive of sympathetic vasomotor tones are unknown. During the current grant period we have found that blockade receptors for excitatory amino acid neurotransmitters (EAA) in the RVLM will reduce blood pressure to the same extend as total autonomic blockade provide that neurons in the caudal ventrolateral medulla are inhibited. Based primarily on this observation, we have developed the hypothesis that tonically active EAA-mediated inputs to the RVLM excite RVLM-sympathoexcitatory neurons and simultaneously excite inhibitory inputs to these neurons via a circuit through the caudal ventrolateral medulla. Furthermore, we hypothesize that this balance between excitatory and inhibitory inputs to RVLM- sympathoexcitatory neurons controlled tonically by EAA-mediated inputs to the RVLM governs the long-term control of sympathetic vasomoter tone, and alteration of this balance may result in hypertension. Furthermore, based on preliminary data we proposed that the tonically- active EAA-mediated input to the RVLM comes from a specific region of the pontine reticular formation. To test these hypothesis, we propose a series of experiments to be conducted in anesthetized as well as conscious rats. Most experiments involved recording blood pressure and sympathetic nerve activity while altering the function of discrete regions of the brain stem by microinjection of neuroactive drugs. Five specific aims will be addressed: [1] to further examine the role of EAA-mediated inputs to the RVLM in the tonic regulation of sympathetic vasomotor tone; [2] to determine whether tonically-active EAA-mediated inputs to the RVLM originate from the pontine reticular formation; [3] to determine if the effects of tonically-active EAA-mediated inputs to the RVLM are altered in the chronic absence of baroreceptor feedback to the brain; [4] to determine whether the balance between excitatory and inhibitory inputs to RVLM sympathoexcitory neuron driven by tonically- active EAA-mediated inputs to the RVLM is altered in models of experimental hypertension; and [5] to begin to determine the role of the C1 population of neurons in the RVLM in mediating the responses driven by tonically-active inputs to the RVLM. These studies will contribute to our understanding of the neural control of blood pressure, and therefore may provide new insight to the pathogenesis and treatment of hypertension.

This Core will supervise purchasing, housing, and behavioral testing of animals. In addition, 6-OHDA-induced lesions will be conducted under the supervision of the Core Director. Additional surgical procedures of general importance to the Program Project also will be developed and performed through the Core as the need arises. In a series of studies we will examine an alternative to the conventional mode of lesioning by employing an osmotic minipump to administer the 6-OHDA. The quality of the surgical work will be monitored via analyses conducted by the neurochemistry and histology services (Cores B and C). This Core should therefore greatly enhance the productivity of the Program, permitting us to integrate the results obtained by different components of the Program Project.

[unreadable] DESCRIPTION (provided by applicant): This application requests funds to continue our integrated basic neuroscience training program at the University of Pittsburgh and Carnegie Mellon University. This training grant (T32 NS07433), currently in its fourth year of funding, has been successful in recruiting and training high quality predoctoral students in neuroscience. Funds are requested to support first and second year graduate students in the Center for Neuroscience at the University of Pittsburgh and the Center for the Neural Basis of Cognition, which is a joint center of the University of Pittsburgh and Carnegie Mellon University, with support being provided to the top one-third of our trainees. The program described in this application focuses primarily on research training in the laboratories of a large and diverse neuroscience training faculty. Students begin laboratory research immediately upon entering the program, and typically rotate through two laboratories in their first year. The training faculty, consisting of 68 faculty from the University of Pittsburgh and Carnegie Mellon University, provides expertise in neuroscience ranging from cellular and molecular to developmental to systems to perception and cognition, and students are exposed to this breadth of neuroscience. In addition to research, students take a series of two intensive one-term core courses in basic neuroscience, at least three elective courses in neuroscience or related areas, a course in statistics, and a variety of seminars and journal clubs. Trainees also participate actively in a series of professional development workshops that provide explicit training in such "survival skills" as written and oral communication, obtaining jobs and grants, teaching, and managing a research lab. Training in the responsible scientific conduct is an integral part of the professional development workshops, the core curriculum, and laboratory training. Students are encouraged to consider a wide range of employment opportunities within which to exercise their skills in research, and seminars are held to permit them to become familiar with employment both within and outside of traditional academic research universities. A solid structure is in place to mentor the trainees and monitor their progress through the program. Students pass through a series of milestones, including the first-year Reprint Exam, a second-year research evaluation, and a grant proposal-based comprehensive exam in the third year, prior to submitting a thesis proposal and progressing to full-time thesis research. Each student has an advising committee to see them through these milestones and assist with their mentoring. Data are presented to document that we recruit outstanding trainees and provide them with excellent training. [unreadable] [unreadable]

DESCRIPTION (provided by applicant): This application requests funds to continue our integrated basic neuroscience training program at the University of Pittsburgh and Carnegie Mellon University. This training grant (T32 NS07433), currently in its ninth year of funding, has been successful in recruiting and training high quality predoctoral students in neuroscience. Funds are requested to support eight first and second year graduate students in the Center for Neuroscience at the University of Pittsburgh and the Center for the Neural Basis of Cognition, which is a joint center of the University of Pittsburgh and Carnegie Mellon University, with support being provided to the top quarter of our trainees. The program described in this application focuses primarily on research training in the laboratories of a large and diverse neuroscience training faculty. Students begin laboratory research immediately upon entering the program, and do two or three semester-long laboratory research rotations in their first year. The training faculty, consisting of 73 faculty from the University of Pittsburgh and Carnegie Mellon University, provides expertise in neuroscience ranging from cellular and molecular to developmental to systems and cognition, and students are exposed to this breadth of neuroscience. In addition to research, students take a series of two intensive one-term core courses in basic neuroscience, at least three elective courses, a course in statistics, and a variety of seminars and journal clubs. Trainees also participate in a series of professional development workshops that provide explicit training in such "survival skills" as written and oral communication, obtaining jobs and grants, teaching, and managing a research lab. Training in the responsible scientific conduct is an integral part of the professional development workshops, the core curriculum, and laboratory training. A solid structure is in place to mentor the trainees and monitor their progress through the program. Students pass through a series of milestones, including the first-year Reprint Exam, a second-year research evaluation, and a grant proposal-based comprehensive exam in the third year, prior to submitting a thesis proposal and progressing to full-time thesis research. Each student has an advising committee to see them through these milestones and assist with their mentoring. Data are presented to document that we recruit outstanding trainees and provide them with excellent training.

This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5).
The nervous system exerts profound regulatory influence over peripheral physiology and this control is essential for homeostasis and coordinating adaptive changes across physiological systems. The regulatory oversight exerted by the brain necessary for the integrated action within and among these systems is considerable, and the brain meets this challenge through regulation of the autonomic nervous system and neuroendocrine control over the pituitary gland; the general goal of this research program is to understand the organization of this regulation. During the past 25 years much has been learned about how the brain controls the autonomic nervous system and this project continues that work by addressing two fundamental questions that remain unanswered. First, what is the nature of the brain circuitry that allows both selective control of specific tissues and also global control involved in activating all sympathetic outflows as occurs in response to certain stressors. Second, what are the brain circuits that provide for the coordinated control of sympathetic and parasympathetic outflows such that when sympathetic activity is increased parasympathetic activity decreases. This project examines these issues using novel neuroanatomical techniques relying on a genetically engineered virus that gets passed among connected nerve cells thereby allowing the visualization of neural circuits in rats. Understanding the nature of these neural circuits is essential for understanding of physiological regulation in mammals. In addition to addressing key questions pertaining to the organization of central autonomic circuits, these studies will provide strong foundational support for this methodology, which can be generally applied to complex neuroanatomical issues. Also, undergraduate student researchers are incorporated into all aspects of this research program, and this project provides an excellent opportunity to immerse undergraduate students in basic neurobiological research. One component of this is the development of a new course in autonomic neuroscience, in which students will be given their own component of this project to develop, analyze, and present their findings.

Research on the addictive properties of cigarettes has, for many years, focused on nicotine (NIC);studies on human subjects and experimental animals have provided compelling evidence that NIC contributes to smoking. However, it is now clear that other chemicals among the more than 4000 compounds of cigarette smoke also contribute to the addictive properties of cigarettes, at least partly by interacting with the effects of NIC on the brain. Of the many compounds present in cigarette smoke, less than a dozen of these have been identified that interact in significant ways with NIC. These compounds can be viewed as fitting into four major classes: a group of so-called minor alkaloids (e.g., anabasine, nornicotine), the p-carbolines Harman and norharman, acetaldehyde, and additional compounds that inhibit monoamine oxidase (MAO). The goal of this project is to explore, in a well characterized rat model of NIC self-administration, the reinforcing effects of NIC within the context of other components of cigarette smoke that likely contribute to its abuse liability. There are four Specific Aims: [1] To determine the effects of a cocktail of tobacco constituents on the doseresponse relationship for NIC self-administration. Hypothesis: a cocktail of tobacco constituents including the minor alkaloids, p-carbolines, acetaldehyde, and MAO inhibitors will shift the dose-response curve for NIC self-administration, such that NIC is more potent and effective in supporting self-administration. [2] To determine whether changing the concentration of the components of the cocktail of tobacco constituents will alter the acquisition and maintenance of NIC self-administration. Hypothesis: increasing the concentrations of the compounds in a cocktail of tobacco constituents will increase self-administration of a specific dose of IC whereas decreasing the concentrations will decrease self-administration. [3] To determine whether changing the concentrations of compounds in the cocktail of tobacco constituents can offset or promote the reduction in NIC self-administration caused by reducing the dose of NIC over time. Hypothesis: the reduction in NIC self-administration caused by reducing the dose of NIC can be enhanced by also reducing the concentration of the cocktail constituents whereas the effects of reducing the dose of NIC can be lessened (or even prevented) by increasing the concentration of these compounds. [4] To determine the relative importance of the different classes of components in the cocktail in the interactions between NIC and the cocktail of tobacco constituents in supporting self-administration behavior.

Research on the addictive properties of cigarettes has, for many years, focused on nicotine (NIC); studies on human subjects and experimental animals have provided compelling evidence that NIC contributes to smoking. However, it is now clear that other chemicals among the more than 4000 compounds of cigarette smoke also contribute to the addictive properties of cigarettes, at least partly by interacting with the effects of NIC on the brain. Of the many compounds present in cigarette smoke, less than a dozen of these have been identified that interact in significant ways with NIC. These compounds can be viewed as fitting into four major classes: a group of so-called minor alkaloids (e.g., anabasine, nornicotine), the p-carbolines Harman and norharman, acetaldehyde, and additional compounds that inhibit monoamine oxidase (MAO). The goal of this project is to explore, in a well characterized rat model of NIC self-administration, the reinforcing effects of NIC within the context of other components of cigarette smoke that likely contribute to its abuse liability. There are four Specific Aims: [1] To determine the effects of a cocktail of tobacco constituents on the doseresponse relationship for NIC self-administration. Hypothesis: a cocktail of tobacco constituents including the minor alkaloids, p-carbolines, acetaldehyde, and MAO inhibitors will shift the dose-response curve for NIC self-administration, such that NIC is more potent and effective in supporting self-administration. [2] To determine whether changing the concentration of the components of the cocktail of tobacco constituents will alter the acquisition and maintenance of NIC self-administration. Hypothesis: increasing the concentrations of the compounds in a cocktail of tobacco constituents will increase self-administration of a specific dose of IC whereas decreasing the concentrations will decrease self-administration. [3] To determine whether changing the concentrations of compounds in the cocktail of tobacco constituents can offset or promote the reduction in NIC self-administration caused by reducing the dose of NIC over time. Hypothesis: the reduction in NIC self-administration caused by reducing the dose of NIC can be enhanced by also reducing the concentration of the cocktail constituents whereas the effects of reducing the dose of NIC can be lessened (or even prevented) by increasing the concentration of these compounds. [4] To determine the relative importance of the different classes of components in the cocktail in the interactions between NIC and the cocktail of tobacco constituents in supporting self-administration behavior.

This application requests funds to continue our integrated basic neuroscience training program at the University of Pittsburgh. This training grant (T32 NS07433), currently in its 19th year of funding, has been successful in recruiting and training high quality predoctoral students in neuroscience. Funds are requested to support 8 graduate students in their first or second year (typically 2nd year) in the Center for Neuroscience at the University of Pittsburgh (CNUP); these 8 students represent the top ~25% of eligible trainees in the CNUP doctoral program. The program described in this application focuses primarily on research training in the laboratories of a large and diverse neuroscience training faculty. Students begin laboratory research immediately upon entering the program, and rotate through at least two laboratories, for one term each, in their first year. The training faculty, consisting of 76 faculty from the CNUP, provides expertise in neuroscience ranging from cellular and molecular to developmental to systems to perception and cognition, and students are exposed to this breadth of neuroscience. In addition to research, students take a series of two intensive one-term core courses in basic neuroscience, at least three elective courses in neuroscience or related areas, a course in experimental design and data analysis, a course in grant writing, and a variety of seminars and journal clubs. Trainees also participate actively in professional development workshops that provide explicit training in professional skills including written and oral communication, obtaining jobs and grants, teaching, and managing a research lab. Training in the responsible scientific conduct is an integral part of the core curriculum and laboratory training, and is also covered in a course on research ethics for first year students. Students are encouraged to consider a wide range of employment opportunities within which to exercise their skills in research, and a variety of mechanisms permit them to become familiar with employment both within and outside of traditional academic research universities. A solid structure is in place to mentor the trainees and monitor their progress through the program. Students pass through a series of milestones, including the first-year Reprint Exam, a second-year research evaluation, and a grant proposal-based Comprehensive Exam in the third year, prior to submitting a thesis proposal and progressing to full-time thesis research. Each student has an advising committee to see them through these milestones and assist with their mentoring. Data are presented to document that we recruit outstanding trainees and provide them with excellent training.

Project Summary Adolescents appear to be particularly vulnerable to begin using tobacco and other nicotine (NIC)-containing products. NIC exposure during adolescence may have long-lasting effects on adult behavior, promoting continued use of NIC-containing products in adulthood. This is especially important in the context of a potential tobacco regulatory policy limiting NIC content in tobacco products. Will adolescent NIC use impact how adults will respond to low NIC products? Will adolescent use of very low doses of NIC minimize the impact of adolescent exposure to NIC on adult NIC use? Alternatively, might it promote the use of NIC products in adulthood? The goal of the experiments presented in this proposal is to determine, using a rodent model of NIC self-administration (SA), the impact of NIC use during adolescence on the use of very low NIC content (VLNC) cigarettes in adulthood. Embedded in this is whether or not adolescent use of NIC-containing products serves as a gateway to smoking and whether NIC reduction could minimize this risk. The current R21 application is designed around two specific aims. Aim 1 tests the hypothesis that NIC SA during adolescence (either low or standard dose of NIC) increases adult NIC SA, especially at low doses of NIC. To address this question, adolescent male and female rats will be allowed to self-administer a low or a more standard dose of NIC starting in early adolescence and then the NIC dose will be changed when they are adults. The key measure will be amount of NIC SA in adulthood, and how that is altered by different doses of adolescent NIC SA. Aim 2 tests the hypothesis that NIC SA during adolescence (either low or standard dose) increases the motivation to seek NIC in adulthood. This hypothesis will be tested at the end of Aim 1 by examining responding for NIC on a progressive ratio schedule of reinforcement in adult rats that self-administered NIC throughout adolescence and into adulthood. An important component of these experiments is that NIC will be self-administered along with a mildly reinforcing environmental stimulus, to incorporate the reinforcement enhancement actions of NIC, which are an essential behavioral component driving NIC use. These experiments will help understand the potential long-term consequences of adolescent NIC exposure on continued use of NIC-containing products and will have important implications in the context of potential NIC reduction tobacco regulatory policy. This proposal fits with the RFA Research Objective focused on addiction and the characteristics of tobacco products that may impact addiction as well as understanding the behaviors related to tobacco product use. Here we focus on the adolescent exposure to NIC-containing products and on other salient stimuli coupled with NIC delivery in supporting NIC SA or motivation to obtain NIC.

This application requests funds to continue our integrated basic neuroscience training program at the University of Pittsburgh. This training grant (T32 NS07433), currently in its 19th year of funding, has been successful in recruiting and training high quality predoctoral students in neuroscience. Funds are requested to support 8 graduate students in their first or second year (typically 2nd year) in the Center for Neuroscience at the University of Pittsburgh (CNUP); these 8 students represent the top ~25% of eligible trainees in the CNUP doctoral program. The program described in this application focuses primarily on research training in the laboratories of a large and diverse neuroscience training faculty. Students begin laboratory research immediately upon entering the program, and rotate through at least two laboratories, for one term each, in their first year. The training faculty, consisting of 76 faculty from the CNUP, provides expertise in neuroscience ranging from cellular and molecular to developmental to systems to perception and cognition, and students are exposed to this breadth of neuroscience. In addition to research, students take a series of two intensive one-term core courses in basic neuroscience, at least three elective courses in neuroscience or related areas, a course in experimental design and data analysis, a course in grant writing, and a variety of seminars and journal clubs. Trainees also participate actively in professional development workshops that provide explicit training in professional skills including written and oral communication, obtaining jobs and grants, teaching, and managing a research lab. Training in the responsible scientific conduct is an integral part of the core curriculum and laboratory training, and is also covered in a course on research ethics for first year students. Students are encouraged to consider a wide range of employment opportunities within which to exercise their skills in research, and a variety of mechanisms permit them to become familiar with employment both within and outside of traditional academic research universities. A solid structure is in place to mentor the trainees and monitor their progress through the program. Students pass through a series of milestones, including the first-year Reprint Exam, a second-year research evaluation, and a grant proposal-based Comprehensive Exam in the third year, prior to submitting a thesis proposal and progressing to full-time thesis research. Each student has an advising committee to see them through these milestones and assist with their mentoring. Data are presented to document that we recruit outstanding trainees and provide them with excellent training.

This application requests funds to continue our integrated basic neuroscience training program at the University of Pittsburgh. This training grant (T32 NS07433), currently in its 19th year of funding, has been successful in recruiting and training high quality predoctoral students in neuroscience. Funds are requested to support 8 graduate students in their first or second year (typically 2nd year) in the Center for Neuroscience at the University of Pittsburgh (CNUP); these 8 students represent the top ~25% of eligible trainees in the CNUP doctoral program. The program described in this application focuses primarily on research training in the laboratories of a large and diverse neuroscience training faculty. Students begin laboratory research immediately upon entering the program, and rotate through at least two laboratories, for one term each, in their first year. The training faculty, consisting of 76 faculty from the CNUP, provides expertise in neuroscience ranging from cellular and molecular to developmental to systems to perception and cognition, and students are exposed to this breadth of neuroscience. In addition to research, students take a series of two intensive one-term core courses in basic neuroscience, at least three elective courses in neuroscience or related areas, a course in experimental design and data analysis, a course in grant writing, and a variety of seminars and journal clubs. Trainees also participate actively in professional development workshops that provide explicit training in professional skills including written and oral communication, obtaining jobs and grants, teaching, and managing a research lab. Training in the responsible scientific conduct is an integral part of the core curriculum and laboratory training, and is also covered in a course on research ethics for first year students. Students are encouraged to consider a wide range of employment opportunities within which to exercise their skills in research, and a variety of mechanisms permit them to become familiar with employment both within and outside of traditional academic research universities. A solid structure is in place to mentor the trainees and monitor their progress through the program. Students pass through a series of milestones, including the first-year Reprint Exam, a second-year research evaluation, and a grant proposal-based Comprehensive Exam in the third year, prior to submitting a thesis proposal and progressing to full-time thesis research. Each student has an advising committee to see them through these milestones and assist with their mentoring. Data are presented to document that we recruit outstanding trainees and provide them with excellent training.