Gene Therapy Reduces Drinking in Rats with Genetic Predisposition to “Alcoholism”
As a follow up to previous work showing that gene therapy can reduce drinking in rats trained to prefer alcohol, scientists at the U.S. Department of Energy’s Brookhaven National Laboratory have used the same technique to cut drinking in rats with a genetic predisposition for heavy alcohol consumption. The findings, along with additional results on the effects of long-term ethanol consumption on certain aspects of brain chemistry, are published in the May 2004 issue of Alcoholism Clinical and Experimental Research.
“Though we are still early in the process, these results improve our understanding of the mechanism or mechanisms of alcohol addiction and strengthen our hope that this treatment approach might one day help people addicted to alcohol,” said Panayotis (Peter) Thanos, who lead the study in Brookhaven Lab’s medical department.
Genetically predisposed alcohol-preferring rats are a much better model for human alcoholism than the rats used previously, which the scientists had to train to prefer alcohol. Without any training, the genetic alcohol-preferring rats drink, on average, more than five grams of ethanol per kilogram of body weight per day when given a free choice between alcohol and plain water. Genetically non-preferring rats, in contrast, typically consume less than one gram of ethanol per kilogram of body weight per day.
In this study, both groups were treated with gene transfer to increase the level of a brain receptor for dopamine, a chemical important for transmitting feelings of pleasure and reward and known to play a role in addiction. After the gene treatment, the alcohol-preferring rats exhibited a 37 percent reduction in their preference for alcohol and cut their total alcohol consumption in half — from 2.7 grams per kilogram of body weight before treatment to 1.3g/kg after. Non-preferring rats also reduced their drinking preference and intake after gene treatment, but not in nearly as dramatic a fashion. The greatest reductions in alcohol preference and consumption were observed within the first few days after gene treatment, and both preference and consumption returned to pre-treatment levels by day 20.
The gene administered was for the dopamine D2 receptor, a protein shown in various studies to be relevant to alcohol and drug abuse. For example, low levels of dopamine D2 receptors in the brain have been postulated to lead to a reward deficiency syndrome that predisposes certain people to addictive behaviors, including drug and alcohol abuse. The alcohol-preferring rats used in this study have about 20-25 percent lower levels of dopamine D2 receptors when compared to the non-preferring rats, which may, in part, explain their tendency toward heavy drinking.
The scientists delivered the gene by first inserting it into a virus that had been rendered harmless. They then injected the virus directly into the rats’ nucleus accumbens, the brain’s pleasure center. The idea behind this type of gene therapy is to use the virus as a vector to carry the gene to the brain cells, which can then use the genetic instructions to make the D2 receptor protein themselves.
As an additional measure in this study, the scientists used micro-positron emission tomography (microPET) imaging to non-invasively assess the effects of chronic alcohol consumption on D2 receptor levels in alcohol-preferring and non-preferring rats. They measured D2 levels seven weeks after the gene therapy treatment (well after the effects of gene therapy had worn off). D2 receptor levels in alcohol-preferring rats were significantly lower (about 16 percent) compared to that in non-preferring rats. These levels were similar to previous data in naïve preferring and non-preferring rats.
In future studies, the D2 connection to alcoholism will be examined in transgenic mice that are totally depleted of D2. In addition, the scientists plan to develop a second generation D2 vector approach that will provide a longer period of treatment.
“These findings further support our hypothesis that high levels of D2 are causally associated with a reduction in alcohol drinking and may serve as a protective factor against alcoholism,” Thanos said.
This study was funded by the Office of Biological and Environmental Research within the Department of Energy’s Office of Science and by the National Institute of Alcohol Abuse and Alcoholism within the National Institutes of Health.
Scientists at the U.S. Department of Energy’s Brookhaven National Laboratory have shown that increasing the level of a brain protein important for transmitting pleasure signals can turn rats that prefer alcohol into light drinkers, and those with no preference into near teetotalers. The findings, published in the first September 2001 issue of the Journal of Neurochemistry (Vol. 78, No. 5), may have implications for the prevention and treatment of alcoholism in humans.
"This is a preliminary study, but when you see a rat that chooses to drink 80 to 90 percent of its daily fluid as alcohol, and then three days later it’s down to 20 percent, that’s a dramatic drop in alcohol intake - a very clear change in behavior," said Panayotis Thanos, the lead researcher. "This gives us great hope that we can refine this treatment for future clinical use."
Gene Therapy Reduces Drinking in "Alcoholic" Rats
The protein in question is the so-called D2 receptor for dopamine, a chemical that transmits brain signals necessary for experiencing feelings of pleasure and reward. Without receptors for dopamine, the signals get "jammed," and the pleasure response is blunted.
Previous studies have shown that alcohol abuse and other addictive drugs increase the brain’s production of dopamine. But, over time, these drugs also deplete the brain’s D2 receptors. This research has suggested that alcoholics increase their intake to try to override the blunted pleasure response, and that people with low levels of D2 receptors may be predisposed to alcohol abuse. These ideas led the Brookhaven researchers to hypothesize that increasing the level of D2 receptors might decrease alcohol intake.
The researchers tested this hypothesis in experimental rats by injecting a virus that had been rendered harmless and altered to carry the D2 receptor gene directly into the rats’ brains. The idea behind this gene therapy is that the virus acts as a vector or mechanism to deliver the gene to the brain cells in the nucleus accumbens, the brain’s pleasure center, so the cells can make the receptor protein themselves.
To see if the D2 receptor levels actually did increase, the scientists studied the brains of one group of rats using sophisticated imaging techniques. They used a radiotracer, a signal-emitting chemical designed to bind to the D2 receptor protein, then detected the signals in brain images called autoradiographs. The strength of the signals indicated that rats injected with the D2 gene did have higher levels of D2 receptors. The levels peaked three to four days after injection and gradually returned to near baseline after eight days.
Then the scientists examined how the injected genes affected the drinking behavior of rats that had been previously trained to self-administer alcohol. Rats that showed a preference for alcohol over water during training were analyzed separately from those that had no preference.
Among the rats that initially preferred alcohol, those that had received the D2 gene showed a 43 percent drop in their preference for alcohol and drank 64 percent less alcohol than rats that received only a placebo virus with no genes. Even the rats that initially had a low preference for alcohol showed significant reductions in both their preference for and intake of alcohol after treatment with the D2 gene.
"This is the first evidence that overproduction of D2 receptors reduces alcohol intake and suggests that high levels of D2 may be protective against alcohol abuse in humans," Thanos said.
The reduction in drinking preference and behavior in both groups was transient, with both measures returning to baseline levels by eight days after treatment. But a second treatment with the D2 genes produced the same dramatic effect.
"This is just a first step," said Thanos, who is working with Brookhaven biologist Paul Freimuth to produce a better gene-delivery system that will have a longer-lasting effect.
This work was funded by the National Institute for Alcohol Abuse and Alcoholism and the U.S. Department of Energy, which supports basic research in a variety of scientific fields.
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