The Effect of Chronic Morphine Use on the Connections of the D2-enriched Medium Spiny Neurons of the Striatum.
Abstract
To date, there have been no observations of the effect of chronic opiates such as morphine on the functional connectivity of the Medium Spiny Neurons (MSNs), which are critical mediators of the response of the mesolimbic dopaminergic pathway to drugs of abuse. We propose that chronic morphine administration will alter the connectivity of these striatal neurons. Using a pseudorabies virus that is retrogradely transported and changes color from red to green when encountering cre-recombinase, we have traced the connections of the indirect D2-enriched MSNs in mice expressing cre-recombinase in D2 neurons. We propose, and will test, that prolonged morphine, followed by a period of withdrawal, will change the connections of the indirect, D2-enriched pathway. This could help further elucidate the mechanistic changes that occur during chronic drug use and withdrawal.
Introduction
Chronic opioid exposure alters neuronal connectivity in rodents through modifications in dendritic spine formation (Robinson and Kolb, 1999; Robinson et al., 2002). Similar drug-induced changes in connectivity have been found in specific brain regions of heroin addicts (Ma, N., et al., 2010; Ma, N., et al., 2010) and in direct but not indirect medium spiny neurons (MSNs) following experimenter-delivered cocaine (Kim, J., et al., 2011). However, there have been no such studies regarding the effect of chronic opiates, such as morphine, on the functional connectivity of these MSNs. We hypothesized that chronic morphine administration would alter the connectivity of these specific striatal neurons. Connectivity changes in the brain can help explain the lasting effects of chronic drug use and addiction. Once brain chemistry is altered, and an individual is no longer functioning ‘normally’, the changes show a behavioral reinforcement of drug-taking habits. This has major implications for addiction and chronic drug use.
The addiction profile consists of several distinct stages, including acquisition, habituation, and withdrawal. The direct and indirect dopamine striatal pathways seem to have an effect on different stages of addiction (Lobo and Nestler, 2011). In order to understand more comprehensively how the mesolimbic system adapts to chronic drug exposure, we aim to study both pathways. For this study, we focused on the indirect D2- enriched MSNs. D2-neurons function as part of the indirect pathway, which has been shown to be involved in the withdrawal stage of addiction with regards to chronic drug use (Lobo and Nestler, 2011). We used a pseudorabies 263 virus (PRV 263), on animals treated with either chronic doses of morphine, or saline, to trace the connections of these MSNs. The pseudorabies virus contains the brainbow cassette which changes color from red to green when expressed in Cre-containing cells (Lichtman, et al., 2007). This virus is retrogradely transported and, as it is replication competent, will trace multi-order connections (Card, et al., 2011). PRV 263 allows us to directly label the pathway through the help of multi-fluorescence capabilities. The virus is used with transgenic mice, engineered to have the D2 gene linked to cre- recombinase.
Our data showed a significant difference between morphine-treated and control animals in two specific areas: the dentate gyrus and the ventral tegmental area (VTA). On a very basic level, the changes associated with this difference happen in the brain through drug-induced neuronal plasticity. By doing studies such as this one and distinguishing exactly which areas, and in exactly what ways these changes are occurring, we may be better equipped to treat addiction and chronic drug use further down the line.