To learn how drugs promote abuse and produce addiction, researchers focus on animal behaviors that parallel human drug-related behaviors.

By Sue Young-Wilson, NIDA Notes Contributing Writer

Research using animals has contributed immensely to our understanding of drug abuse and its consequences, prevention, and treatment. Animal studies have yielded fundamental insights into why people abuse drugs and how drugs cause the compulsion and disordered thinking seen in addiction. They provide clues to strategies for preventing and treating abuse and addiction. They also provide the first tests of the safety and efficacy of potential new vaccines and medications. Because animal research is so crucial, we have chosen it as the topic for this inaugural article in a new NIDA Notes Reference Series. Each article in the series will discuss a basic idea or theme in drug abuse science, citing NIDA Notes articles as illustrative references.

WHY EXPERIMENT WITH ANIMALS?

Researchers rely on experiments with animals to answer questions about how drugs and potential medications may affect the brain and body without exposing people to potential toxicity, risk of addiction, or invasive medical procedures. These experiments reduce or eliminate many factors that can make studies with people hard to interpret, such as differences in diet, health, drug history, genetic makeup, socioeconomic circumstances, understanding, and attitudes. While results from animal studies should be extrapolated to humans with caution, their value is incalculable. Researchers follow strict ethical guidelines to minimize animals' discomfort and ensure that any adversity animals suffer is unavoidable and warranted in terms of the perceived importance of the knowledge to be gained.

ANIMALS AND ADDICTION

Addiction is a human disease; only people normally have sufficient access to drugs to become addicted, and only people can report some of the hallmark experiences of the disorder, such as craving a substance or trying and failing to stop using it. Yet when laboratory animals (rats, mice, monkeys, baboons, and some other species) are exposed to addictive substances in controlled settings, they manifest equivalents of several of the behaviors we use to recognize and define abuse and addiction. They generally will:

• Take more and more of it,
• Devote much time and energy to getting it,
• Continue taking it despite adverse consequences,
• Undergo a withdrawal syndrome upon sudden cessation, and
• Relapse in response to environmental or stressful triggers or renewed exposure.

Researchers call these behaviors "animal models" of the corresponding aspects of human addictive behavior. The core procedures of animal research on addiction are a set of protocols that assess a substance's addictive properties by exposing animals to it and observing whether they exhibit the model behaviors. Each protocol focuses on one behavior.

THE CORE PROTOCOLS

The animal models of addiction enable researchers to translate questions about a substance's effects on people into parallel questions about whether animals exposed to the substance will exhibit the stand-in behaviors. For example, the question, "Will this substance provide people with an experience they want to repeat, leading to possible abuse?" becomes, "Once animals are exposed to this substance, will they push a lever to obtain more?"

The protocols for administering substances and documenting the model animal behaviors are standardized so that researchers can interpret and compare results. Quantification of the behaviors is indispensable: Investigators seeking to determine whether a substance is likely to be abused, for example, will use a computer to count the number of times test animals push a lever to self-administer it. Scientific instrument makers have provided specialized equipment for measuring the significant behaviors. Thus, for example, researchers assessing a substance's stimulant effects typically will measure a test animal's locomotor activity by putting it into a cage equipped with infrared sensors that automatically monitor its movements.

The most frequently posed questions about human responses to potentially addictive substances, the corresponding questions about animal behavior, and experimental protocols are as follows:

Does the substance have effects that will motivate people to abuse it?
Researchers have developed two independent protocols, each using a different animal model, to answer this question.

1. Self-administration experiments:
Once an animal has experienced a substance's effects, will it work to obtain more?

To answer this question, researchers first train an animal to obtain rewards—usually food—by some voluntary action, for example, by pushing a lever. They then give the animal an initial injection or infusion of the substance in question and place it in a cage where it can self-administer more doses by the same action. Staying with the example of a lever, two conditions must now be met for the researchers to conclude that the substance motivates self-administration:

• The animal persistently pushes the lever; and
• The animal pushes the drug-linked lever more than it pushes an identical lever that delivers either nothing at all or infusions of a control substance, such as saline solution, that has no rewarding properties. This second condition ensures that the animal's goal in pushing the drug-linked lever is to get the drug and not, for example, just for exercise (in which case it would be expected to push both levers equally).

If test animals' behaviors meet both conditions, the drug is said to be reinforcing, meaning it provides an experience animals seek to repeat. NIDA-funded researchers recently used a self-administration experiment to verify their hypothesis that acetaldehyde, a component in tobacco smoke, contributes to adolescent tobacco addiction.¹

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2. Conditioned place preference experiments:
Once an animal has been exposed to a substance, will it prefer the place where it had this experience to other places where it has not?

For this protocol, researchers utilize a test cage consisting of two chambers with markedly different features: for example, one chamber may be black with a Plexiglas floor and the other white with a wire-mesh floor, or they may have distinctive aromas, lighting, or sounds. A passageway joins the two chambers, with doors that can be opened or closed to confine the animal in either chamber or allow free movement between them.

Preparatory to the experiment, the researchers inject a test animal with the substance under investigation and confine it to one chamber. They may expose the animal once or many times, always putting it in the same chamber; the goal is to train, or condition, the animal to associate the drug experience with that chamber and its distinctive features.

On the day of the experiment, the researchers place the animal, uninjected, in the hallway between the two chambers and monitor its movements. They will conclude that the substance is reinforcing if the animal spends most of its time in the chamber it has learned to associate with the substance. Sometimes animals will instead favor the other chamber—a sign they have an aversion to the substance. As in self-administration experiments, researchers incorporate control conditions in these experiments to eliminate alternative explanations for the animals' behaviors. Although conditioned place preference studies do not directly measure drug reinforcement, their results match those of self-administration studies fairly well.

How strongly does the substance motivate continued use?
How much effort will an animal expend to obtain a dose of the substance?

To answer this, researchers set up self-administration experiments just like those used to establish abuse tendency, but with one modification: They program the lever so that the animal must press it more times to obtain each successive infusion of the substance. Typically, the animal receives an infusion after the first press, the second, fourth, eighth, sixteenth, and so on. The strength of reinforcement is indicated by how far the animal goes in this escalation before the effort required for the next dose outstrips its motivation and it desists. Also measured is the pace of the lever-pressing: the shorter the intervals between presses, the more motivated the animal may be.

Will people be motivated to keep on taking the drug even when they know that negative consequences will result?
Will animals continue to press a lever that delivers the substance, but is accompanied by an unpleasant sensation?

Researchers set up self-administration studies with yet another wrinkle: When the animal presses the lever, it receives, along with the substance, a discomfiting stimulus such as a tail pinch or an electrical shock to its foot. If the animal keeps pressing the lever nonetheless, it is considered likely that people, too, may overvalue the substance in relation to their health and well-being.

Will people who take the drug for long periods experience withdrawal?
Will animals that have been extensively exposed to the substance show signs of discomfort or dysfunction if they abruptly lose access to it?

A person is at risk for withdrawal symptoms when his or her body is dependent upon a substance—that is, has adapted physiologically to the substance and now needs it to function properly. To replicate this state in laboratory experiments, researchers will either directly administer a substance to an animal or place an animal in a cage where it can freely self-administer the substance. In the latter case, the researchers will watch for the animal to settle into a stable rhythm of lever-pressing or to steadily increase its intake of the substance, each of which suggests its body has reached a new physiological equilibrium that accommodates the substance's effects. At this point, the scientists cut off the animal's access to the substance and observe its response. Withdrawal is inferred if the animal develops certain abnormal behaviors. In one such experiment, described in this issue, baboons chronically exposed to the club drug GHB—also known as "liquid ecstasy"—developed tremors, loss of appetite, and agitation when drug administration stopped.²

Will a person who becomes abstinent be at risk for relapse?
Will a triggering stimulus cause an animal to start seeking a drug again after having stopped?

Experiments to test relapse have three basic steps:

• Establishment of dependence: As in studies to test withdrawal, an animal is steadily infused or placed in a cage where it self-administers the substance under investigation until its pattern of lever-pressing indicates dependence;
• Extinction: The researchers leave the animal in the cage but disconnect the lever from the substance delivery mechanism. The animal keeps on pressing the lever for a while, but with no substance forthcoming, eventually stops. In lab parlance, the animal's lever-pressing behavior is now "extinguished," and it is in a state comparable to that of a person who has achieved abstinence;
• Exposure to a relapse trigger: The researchers now expose the animal to a stimulus they believe can trigger relapse. If the animal resumes avid lever-pressing, the researchers conclude that the substance's effects include an abiding potential for relapse.

In relapse experiments, researchers test animals with stimuli corresponding to those that most commonly cause people who are abstaining from drugs to relapse: stress, environmental cues associated with the drug experience, and reexposure to the drug (for example, when someone decides it won't hurt to smoke just one cigarette and quickly is back to a pack a day). An environmental cue often used in animal models of relapse is a light that turns on when the lever is primed to deliver the substance during the self-administration stage, turns off in the extinction stage, and reappears in the last stage. Stress often is induced with mild electrical shocks.³

BEYOND THE CORE PROTOCOLS

Animal studies are useful beyond determining whether substances have addictive properties. They also guide the development of new treatments and provide key information about how drugs produce their addictive and other effects.

Evaluating treatments

To assess whether a vaccine or medication is likely to help prevent or treat abuse of a drug, researchers typically follow the same protocols they use to assess the drug's reinforcement potential, but with two modifications. At an appropriate stage in the experiment, they give some of their animal subjects the potential vaccine or medication, and they use a control group of drug-exposed but untreated or placebo-treated animals.

Investigations into drugs' effects have contributed greatly to our general understanding of how the brain functions in such fundamental areas as learning and memory, motivation, and pain processing.

In an experiment to test a vaccine, researchers ask: "Compared with the control group, will the animals we pretreat with the vaccine before giving them access to the drug self-administer less of it, or demonstrate less preference for the cage in which they were exposed to it?"⁴ In a study to evaluate a potential medication to reduce withdrawal symptoms, they ask: "If animals are physically dependent on the drug, will a group that is treated with the proposed medication show fewer signs of withdrawal than an untreated group when the drug is stopped?" If a medication is being evaluated for efficacy against relapse, they ask: "If we bring animals through the progression of self-administration and extinction and then expose them to a relapse trigger, will the medicated animals revert to avid self-administration less readily than the control group?"⁵

Studies of drug mechanisms

Information on how drugs act on the brain to produce abuse and addiction provides the basis for targeted medication development: Once research shows that a drug of abuse affects a particular brain process, pharmacologists can identify and test medications that have opposing effects. Research into the neurobiological mechanisms of abuse and addiction also sheds light on genetic and physiological traits that heighten or reduce vulnerability to drug abuse. Investigations into drugs' effects have contributed greatly, as well, to our general understanding of how the brain functions in such fundamental areas as learning and memory, motivation, and pain processing.⁶

Researchers learn about drug mechanisms and some effects, such as toxicity, by simply exposing animals to drugs and analyzing the impact on brain structures and systems.⁷ Other studies try to decipher how drugs' biological effects give rise to the behaviors associated with abuse and addiction: Typically, these studies combine the core animal experimental protocols with specialized techniques of pharmacological, neurobiological, or genetic investigation.⁸

THE LIMITS AND FUTURE OF ANIMAL EXPERIMENTATION

Animal experiments shed light primarily on drug responses that are based in the many brain and bodily systems and processes that humans share with other species. Yet animal experiments can never yield a complete picture of human interactions with drugs. While evolution equipped people and animals with the same or very similar mechanisms for many basic aspects of motivation and inhibition, learning and memory, and pain response, the human brain integrates these with speech and writing, logic, and symbolic reasoning. Moreover, only in humans do social factors such as laws, culture, religion, and economics modify the likelihood and consequences of drug abuse. Accordingly, results from animal studies need to be interpreted with caution, and human studies alone can reveal influences on abuse and addiction that involve uniquely human traits and capacities.

Recent technological advances enable today's researchers to investigate some issues directly in humans that previously would have required animal experimentation. The most striking example, perhaps, is the use of noninvasive imaging techniques to monitor the brain's complex responses to drugs while they are occurring. The same techniques also permit new types of experiments in animals.⁹ Other technological advances have extended the use of animal experiments into new fields of investigation; for example, researchers can now investigate the role of a particular gene in drug abuse through the use of specifically bred strains of mice.¹⁰ Researchers' ingenuity continues to yield new ways to exploit the unique advantages of animal models to deepen our knowledge of drug abuse and addiction.

Sources

¹ Study Points to Acetaldehyde-Nicotine Combination in Adolescent Addiction, NIDA Notes, Vol. 20, No. 3.

² Animal Research Shows GHB Acts on GABA Receptors, NIDA Notes, Vol. 20, No. 5.

³ A Single Cocaine "Binge" Can Establish Long-Term Cue-Induced Drug-Seeking in Rats, NIDA Notes, Vol. 19, No. 6.

⁴ Nicotine Vaccine Moves Toward Clinical Trials, NIDA Notes, Vol. 15, No. 5.

⁵ Animal Studies Suggest D3 Receptors Offer New Target for Treatment Medications, NIDA Notes, Vol. 18, No. 4.

⁶ NIDA Research Identifies Proteins That Direct Formation of the Brain's Communication Circuits, NIDA Notes, Vol. 20, No. 3.

⁷ Stimulant Drugs Limit Rats' Brain Response to Experience, NIDA Notes, Vol. 19, No. 3; Novel Cannabinoid Appears Promising for Treatment of Chronic Pain, NIDA Notes, Vol. 19, No. 2.

⁸ Dopamine Enhancement Underlies a Toluene Behavioral Effect, NIDA Notes, Vol. 19, No. 5; Site on Brain Cells Appears Crucial to Nicotine Addiction, NIDA Notes, Vol. 20, No. 2; Researchers Investigate Cocaine "Abstinence Syndrome," NIDA Notes, Vol. 20, No. 1; Brain Glutamate Concentrations Affect Cocaine Seeking, NIDA Notes, Vol. 19, No. 3.

⁹ Social Environment Appears Linked to Biological Changes in Dopamine System, May Influence Vulnerability to Cocaine Addiction, NIDA Notes, Vol. 17, No. 5.

¹⁰ Cocaine's Pleasurable Effects May Involve Multiple Chemical Sites, NIDA Notes, Vol. 14, No. 2.

Volume 20, Number 5 (April 2006)