Animal models of relapse, known as reinstatement procedures, have been used extensively to study the role of stress in relapse to drug seeking. Here, we report on a method for inducing the reinstatement of cocaine seeking in laboratory rats via acute exposures to mild, intermittent electric footshock.
The most insidious aspect of drug addiction is the high propensity for relapse. Animal models of relapse, known as reinstatement procedures, have been used extensively to study the neurobiology and phenomenology of relapse to drug use. Although procedural variations have emerged over the past several decades, the most conventional reinstatement procedures are based on the drug self-administration (SA) model. In this model, an animal is trained to perform an operant response to obtain drug. Subsequently, the behavior is extinguished by withholding response-contingent reinforcement. Reinstatement of drug seeking is then triggered by a discrete event, such as an injection of the training drug, re-exposure to drug-associated cues, or exposure to a stressor 1.
Reinstatement procedures were originally developed to study the ability of acute non-contingent exposure to the training drug to reinstate drug seeking in rats and monkeys 1, 2. Reinstatement procedures have since been modified to study the role of environmental stimuli, including drug-associated cues and exposure to various forms of stress, in relapse to drug seeking 1, 3, 4.
Over the past 15 years, a major focus of the reinstatement literature has been on the role of stress in drug relapse. One of the most commonly used forms of stress for studying this relationship is acute exposures to mild, intermittent, electric footshocks. The ability of footshock stress to induce reinstatement of drug seeking was originally demonstrated by Shaham and colleagues (1995) in rats with a history of intravenous heroin SA5. Subsequently, the effect was generalized to rats with histories of intravenous cocaine, methamphetamine, and nicotine SA, as well as oral ethanol SA 3, 6.
Although footshock-induced reinstatement of drug seeking can be achieved reliably and robustly, it is an effect that tends to be sensitive to certain parametrical variables. These include the arrangement of extinction and reinstatement test sessions, the intensity and duration of footshock stress, and the presence of drug-associated cues during extinction and testing for reinstatement. Here we present a protocol for footshock-induced reinstatement of cocaine seeking that we have used with consistent success to study the relationship between stress and cocaine seeking.
1. Animal Housing and Acclimatization Procedures
2. Intravenous Catheterization Surgery
3. Behavioral Procedures
All behavioral procedures are carried out using equipment supplied by Med Associates Inc. (St Albans, VT). The equipment, housed in a dedicated room, includes a set of standard operant chambers, each contained within a sound-attenuating enclosure. Each operant chamber is equipped with two retractable levers, both elevated 6.5 cm above a stainless steel rod floor. A white stimulus light is located just above one lever, and responses on that lever (the so-called “active” lever) result in the simultaneous illumination of the stimulus light and activation of an infusion pump (Razel Scientific Instruments, St. Albans, VT). The pump is situated on a shelf just outside the operant chamber, but within the sound-attenuating enclosure. The other lever (the so-called “inactive” lever) is left extended in the chamber throughout all experimental procedures; responding on this lever is without consequence. Finally, each chamber is equipped to deliver constant-current, intermittent, inescapable, electric footshock through a scrambler to the steel rod floor. All experimental conditions (e.g., stimulus presentations, drug infusions, shock delivery), as well as data recording and consolidation, are accomplished via interfaces between the test chambers and a central computer operating Med Associates software (MED-IV).
The experiment is carried out in 5 phases: 1) Habituation; 2) Cocaine SA; 3) Drug-free period; 4) Extinction, and 5) Testing for reinstatement. In all phases (except for the drug-free period), rats are transferred for daily sessions from the housing room to the operant test chambers. Animals are transferred individually, in small plastic containers. Food and water are not available in the operant chambers.
4. Representative Results:
Figure 1. Stylized response profiles for activity on the active and inactive levers during self-administration training, extinction, and reinstatement testing.
Figure 1 presents stylized operant response profiles, typical of those obtained during the various phases of the experimental protocol described herein.
1. Active lever responses
2. Inactive lever responses
It can be seen in Figure 1 (red line) that, throughout all experimental phases, there are very low response levels associated with the inactive lever. However, a modest increase in responding on this lever is sometimes observed following exposure to footshock stress. Even in cases in which such an increase in responding occurs, the increase in active lever responding tends to be proportionately much greater.
Over the past few decades, three variations of the SA reinstatement procedure have been developed to study reinstatement of drug seeking in laboratory rats and monkeys. The differences that define the procedural variations relate primarily to whether drug SA, extinction, and testing sessions occur within the same day or on different days 1. The procedure we have described in this paper is commonly referred to as the “within-between” session variation, because drug SA occurs during daily sessions (i.e., in a between-session manner) that are separate from extinction and test sessions, but extinction and testing occur on the same day (i.e., in a within-session manner) [e.g., 7, 9]. Alternatively, in so-called “between sessions” procedures, all phases of the experiment occur sequentially, such that SA, extinction, and testing sessions are given on different days (i.e., all phases are between sessions) [e.g., 8, 10, 11]. In contrast, in complete “within sessions” procedures, SA training, extinction, and test sessions all occur within the same day [e.g., 12].
Although there are published reports of footshock-induced reinstatement of drug seeking using different variations of the reinstatement procedure, we have found this stressor (at least in animals with a history of cocaine SA) to be most effective when extinction and testing occur on the same day (i.e., in a “within-between” sessions manner). Alternatively, we have had success with obtaining reliable footshock-induced reinstatement of cocaine seeking using a completely “between sessions” procedure, but only under conditions in which animals are housed in the SA chambers throughout the experiment 8, or at least during extinction and testing 13. The disadvantage of this between-session approach, over that described in the present protocol, is that it limits the number of subjects that can be trained and tested at one time and, thereby, decreases the efficiency of data collection. Although under the conditions of the present protocol only one group of animals can be extinguished and tested at a given time, a greater throughput can be achieved by staggering by one week the start time of the SA training phase for two groups of rats. In this scenario, a first group of rats can enter the drug-free phase as the second group enters Week 2 of SA training, and, subsequently, extinction and testing can be carried out in the first group while the second group is in its drug-free phase. This approach permits a more efficient use of resources and accelerates the rate of data acquisition, while leading to very effective and reliable footshock-induced reinstatement.
Importantly, in our procedure, extinction of the previously drug-reinforced behavior and testing for reinstatement occurs in the presence of drug-associated cues. Although we have never carried out extinction training or testing in the absence of drug-associated cues, it is our understanding, based on personal communications, that the stressor may in fact be ineffective in inducing reinstatement when testing occurs in the absence of extinguished cues (at least in the case of animals with a history of cocaine SA). A systematic examination of this variable, and the theoretical implications it poses, is an important question for future research.
A final procedural factor that warrants comment is that the effect of footshock stress on reinstatement appears to be highly sensitive to manipulations of context. For example, footshock stress is only effective in inducing the reinstatement of drug seeking if it is administered in the environment in which drug SA and extinction occur; that is, footshock does not reinstate drug seeking if exposure to the stressor occurs in a novel context, suggesting an important interaction between the stressor and drug environment in its effects on reinstatement 14.
In conclusion, footshock reinstatement procedures, like those described in this paper, have been used with considerable success to elucidate the basic behavioral and neurobiological mechanisms governing the relationship between stress and drug relapse. Indeed, these procedures, when modified to accommodate pharmacological pretreatments or neurochemical manipulations targeting specific neuronal substrates, can serve as powerful tools for characterizing the neurobiology of reinstatement to drug seeking by stress.
The authors have nothing to disclose.
Material Name | Typ | Company | Catalogue Number | Comment |
---|---|---|---|---|
Basic Drug Self-Administration Test Package for Rat Package | Med Associates, Inc | MED-008-CT-B1 | ||
Deep extra tall MDF sound attenuating cubicle | Basic Drug Self-Administration Test Package for Rat Package Component | Med Associates, Inc | ENV-018MD | |
Standard modular test chamber with drug infusion top and waste pan | Basic Drug Self-Administration Test Package for Rat Package Component | Med Associates, Inc | ENV-008CT | |
Stainless steel grid floor | Basic Drug Self-Administration Test Package for Rat Package Component | Med Associates, Inc | ENV-005 | |
House light, hooded, 28V | Basic Drug Self-Administration Test Package for Rat Package Component | Med Associates, Inc | ENV-215M | |
Stimulus light 1” white lens | Basic Drug Self-Administration Test Package for Rat Package Component | Med Associates, Inc | ENV-221M | |
Low profile retractable response lever | Basic Drug Self-Administration Test Package for Rat Package Component | Med Associates, Inc | ENV-112CM | |
SmartCtrl 8 Input/16 output package | Basic Drug Self-Administration Test Package for Rat Package Component | Med Associates, Inc | DIG-716P2 | |
Single speed syringe pump, 3.33 RPM | Basic Drug Self-Administration Test Package for Rat Package Component | Med Associates, Inc | PHM-100 | |
Rat self infusion package | Basic Drug Self-Administration Test Package for Rat Package Component | Med Associates, Inc | PHM-119B-23-D | |
Drug Delivery, Arm assembly | Basic Drug Self-Administration Test Package for Rat Package Component | Med Associates, Inc | PHM-110-SAI | |
Square wave aversive stimulator | Footshock Stress Component | Med Associates, Inc | ENV-414S | |
Patented quick disconnect harness for stainless steel grid floor | Footshock Stress Component | Med Associates, Inc | ENV-005-QD | |
Shock output cable | Footshock Stress Component | Med Associates, Inc | SG-219G-10 | |
MED-PC IV Software | Software | Med Associates, Inc | SOF-735 |