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Cardiovascular responses to nicotine

Institution: University of California, San Diego
Investigator(s): Palmer Taylor, Ph.D.
Award Cycle: 1997 (Cycle 6) Grant #: 6RT-0009 Award: $430,843
Subject Area: Nicotine Dependence
Award Type: Research Project Awards
Abstracts

Initial Award Abstract
We propose to extend our initial studies on the receptors and neural pathways through which nicotine and related compounds exert their actions at the level of the spinal cord. The spinal cord integrates sensory stimuli from the periphery through reflexes and manifests these stimuli in autonomic (i.e., cardiovascular) and motor responses. Signals are sent to higher centers of the brain to modulate spinal actions. Substantial evidence suggests that these responses are either transmitted or amplified through acetylcholine acting on spinal nicotinic receptors. These receptors are on nerve endings and amplify the release of other neurotransmitter. Thus, nicotine stimulates the various spinal pathways that control blood pressure, heart rate, and certain sensory responses. The actions of nicotine at the level of the spinal cord mimic the cholinergic amplification and give rise to increased blood pressure, heart rate, and irritation. These responses and their alteration with chronic nicotine use may be associated with cardiovascular disease.

We have approached the action of nicotine in the spinal cord at several levels. The first involves implantation of catheters into the spinal column of rats and injection of nicotine at various segmental levels of the spinal cord. Blood pressure, heart rate and irritation responses are monitored in the rat. Hence this provides a means of monitoring integrated responses of the animal. Cellular responses to nicotine will be monitored by implanting microdialysis tubing in the spinal column and measuring the release of neurotransmitter. The receptors themselves are defined through the specificity of binding of nicotine and related compounds in specified regions of the spinal cord. We have also devised methods to detect and localize the messenger RNA encoding the receptor subunits. This approach, along with the identification of the subunits with specific antibodies, provides a means for localizing the nicotinic receptors and determining their subunit compositions in various regions of the spinal column and in the spinal roots.

Our proposed project, which extends from monitoring responses in the animal to detailed molecular events, should provide a comprehensive analysis of nicotine's action in relation to the relevant pathways in the spinal cord. This study should contribute substantially to understanding the deleterious effects of chronic nicotine use on the cardiovascular system.

Final Report
Spinal nicotinic receptor stimulation results in various physiological responses including cardiovascular, nociceptive as well as antinociceptive responses. Pharmacological studies indicate that these varieties of responses appear to be elicited following stimulation of distinct subtypes of spinal nicotinic receptors. Our goal has been to localize the nicotinic receptors in various spinal neurons and terminals and correlate their location with an already described specific pharmacological response. To achieve this end we have generated polyclonal antibodies to specific neuronal nicotinic receptor subunits to identify the expression of these subunits in various spinal cord regions. In order to localize the expression of the subunit proteins in various neurons and terminal, we have utilized specific pharmacological manipulations to eliminate a specific group of neurons. Accordingly, neonatal capsaicin treatment has been done in rats to eliminate the primary afferent terminals in the dorsal lumbar spinal cord. Neonatal capsaicin treatment significantly reduced CGRP and substance P content in the superficial dorsal horn region in treated as com~ared to vehicle-treated control rats. Receptor autoradiography indicates significant decrease in [ H]-epibatidine binding in the same region. Moreover, nociceptive response to spinal nicotinic agonist like epibatidine is also significantly decreased in the capsaicin-treated versus control rats. Immunohistochemical studies with the polyclonal antibodies against nicotinic receptor subunits indicate that there is significant loss of staining for alpha 3, alpha 5, beta 2 and 04 as well as isolectin B4 and CGRP (markers for primary afferent terminals) in the superficial dorsal spinal cord region in treated versus control rats. These subunits also exhibit colocalization with isolectin B4 and CGRP in the same region. In support to the above observation, the above nicotinic receptor subunits are also found to be expressed in DRG neurons grown in culture. In contrast to the above four nicotinic receptor subunits listed above, a4 subunit is not found to be present on the primary afferent terminals nor expressed in the DRG neurons. However, a4 is expressed in the motor neurons in the ventral spinal cord as well as in motor neuron culture in vitro. Alpha 5 and beta 2 are also expressed in the motor neurons as well. Overall our data indicate that different subtype of nicotinic receptors may be expressed in the motor neurons as compared to that on primary afferent terminals. Our data also indicate that a significant portion of the nociceptive response elicited by spinal nicotinic agonists is mediated via stimulation of neuronal nicotinic receptors present on primary afferent terminals.
Publications

Nociceptive and antinociceptive responses to intrathecally administered nicotinic agonists
Periodical: Neuropharmacology Index Medicus:
Authors: Khan IM, Buerkle H, Taylor P, Taksh TL ART
Yr: 1998 Vol: 37 Nbr: Abs: Pg: 1515-1525