Propranolol Administration During Morphine Addiction Attenuates Reinstatement of Drug-Aversive Memories Caused by Exposure to Stressful Stimuli methods
Aim. Evidence-backed execution summary for Propranolol Administration During Morphine Addiction Attenuates Reinstatement of Drug-Aversive Memories Caused by Exposure to Stressful Stimuli methods from Propranolol Administration During Morphine Addiction Attenuates Reinstatement of Drug-Aversive Memories Caused by Exposure to Stressful Stimuli.
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mouse
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1. Introduction
reagent used in the protocol.
- Use
- Understanding whether stress triggers the retrieval of drug-withdrawal memories through activation of the sympathetic nervous system is important for developing novel therapeutic strategies to prevent relapse. Muscle activity measurements offer a potential readout of such stress responses. In this study, a paradigm...
4. Materials and Methods
reagent used in the protocol.
- Use
- Mice were randomly assigned to one of two treatment conditions: chronic saline ( n = 86) or chronic morphine ( n = 93). With the purpose of inducing dependence on morphine, the drug was administered intraperitoneally once daily in escalating doses: 10 mg/kg on day 1, 30 mg/kg on day 2, 50 mg/kg on day 3, and 60 mg/k...
4.2. Conditioned Place Aversion (CPA)
reagent used in the protocol.
- Use
- CPA expression was evaluated on day 5 under drug-free conditions by using the same procedure as in the pre-test. The time spent in the naloxone-paired chamber was recorded and registered.
4.4. Reinstatement of Conditioned Place Aversion
reagent used in the protocol.
- Use
- Reinstatement refers to the re-emergence of a previously extinguished conditioned response following exposure to an unconditioned stimulus-such as a drug, drug-associated cues, or stressors-when extinction has previously occurred [, ]. In the present study, reinstatement of naloxone-induced CPA was eval...
1. Introduction
Understanding whether stress triggers the retrieval of drug-withdrawal memories through activation of the sympathetic nervous system is important for developing novel therapeutic strategies to prevent relapse. Muscle activity measurements offer a potential readout of such stress responses. In this study, a paradigm...
- Use
- Understanding whether stress triggers the retrieval of drug-withdrawal memories through activation of the sympathetic nervous system is important for developing novel therapeutic strategies to prevent relapse. Muscle activity measurements offer a potential readout of such stress responses. In this study, a paradigm...
4.2. Conditioned Place Aversion (CPA)
CPA expression was evaluated on day 5 under drug-free conditions by using the same procedure as in the pre-test. The time spent in the naloxone-paired chamber was recorded and registered.
- Use
- CPA expression was evaluated on day 5 under drug-free conditions by using the same procedure as in the pre-test. The time spent in the naloxone-paired chamber was recorded and registered.
4.3. Extinction of CPA
Extinction sessions began 24 h after the CPA post-test and were conducted daily for 20-25 days until extinction criteria were met. Extinction was confirmed using paired t-test analyses comparing post-test and final extinction values. The procedure during each session started when the mice were placed in the ce...
- Use
- Extinction sessions began 24 h after the CPA post-test and were conducted daily for 20-25 days until extinction criteria were met. Extinction was confirmed using paired t-test analyses comparing post-test and final extinction values. The procedure during each session started when the mice were placed in the ce...
4.4. Reinstatement of Conditioned Place Aversion
Reinstatement refers to the re-emergence of a previously extinguished conditioned response following exposure to an unconditioned stimulus-such as a drug, drug-associated cues, or stressors-when extinction has previously occurred [, ]. In the present study, reinstatement of naloxone-induced CPA was eval...
- Use
- Reinstatement refers to the re-emergence of a previously extinguished conditioned response following exposure to an unconditioned stimulus-such as a drug, drug-associated cues, or stressors-when extinction has previously occurred [, ]. In the present study, reinstatement of naloxone-induced CPA was eval...
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1. Introduction
Understanding whether stress triggers the retrieval of drug-withdrawal memories through activation of the sympathetic nervous system is important for developing novel therapeutic strategies to prevent relapse. Muscle activity measurements offer a potential readout of such stress responses. In this study, a paradigm to measure the re-emergence of an aversive contextual memory induced by physical (restraint and tail-pinch) and psychosocial (social defeat) stressors was applied in mice previously subjected to the CPA procedure. The aim of this research was to expand knowledge about the role of β-ARs in the retrieval of aversive memories and to assess the potential of β-AR antagonism to mitigate physical and/or psychosocial stress-induced negative memories that contribute to possible relapse.
4. Materials and Methods
Adult male C57BL/6J mice ( n = 179), which weighed 25-30 g at the beginning of the experiment, were obtained from the Animal Facilities of the University of Murcia (CEIB), Murcia, Spain. These animals were divided into groups of 4-6 per cage under controlled temperature conditions, with access ad libitum to food and water. A 12-h light/dark cycle was established throughout the study. Prior to the experimental procedures, mice were handled and exposed to the testing environment for at least one week. All procedures in this study have adhered to the European Communities Council Directive 2010/63/EU (22 September 2010) and were approved by the Animal Ethics Committee of the University of Murcia (CEEA 534/2019).
4. Materials and Methods
Mice were randomly assigned to one of two treatment conditions: chronic saline ( n = 86) or chronic morphine ( n = 93). With the purpose of inducing dependence on morphine, the drug was administered intraperitoneally once daily in escalating doses: 10 mg/kg on day 1, 30 mg/kg on day 2, 50 mg/kg on day 3, and 60 mg/kg on day 4. This dosage has been previously validated to induce opioid tolerance and dependence [,,, ]. Instead, control animals received equivalent volumes of saline following the same schedule. Subsequently, animals were randomly assigned to acute treatment conditions (vehicle or propranolol) and according to stress type (physical, psychosocial, or control group). Behavioral scoring was performed blind to treatment, thereby strengthening the rigor of the experimental design.
4.3. Extinction of CPA
Extinction sessions began 24 h after the CPA post-test and were conducted daily for 20-25 days until extinction criteria were met. Extinction was confirmed using paired t-test analyses comparing post-test and final extinction values. The procedure during each session started when the mice were placed in the central passageway and allowed free access to all chambers for 15 min. Time spent in the naloxone-paired chamber and the number of crossings between compartments were recorded.
4.4. Reinstatement of Conditioned Place Aversion
Reinstatement refers to the re-emergence of a previously extinguished conditioned response following exposure to an unconditioned stimulus-such as a drug, drug-associated cues, or stressors-when extinction has previously occurred [, ]. In the present study, reinstatement of naloxone-induced CPA was evaluated one day after extinction by exposing mice to either an acute social defeat or a physical stressor (restraint or tail-pinch). The reinstatement test followed the same procedure as the pre-test, post-test, and extinction sessions, allowing animals free access to all compartments of the apparatus for 15 min. To minimize contextual associations, all reinstatement procedures were conducted in a room distinct from that used during conditioning and extinction phases, thereby ensuring a non-contingent spatial context relative to the original drug-paired environment.
4.5.2. Experiment 2: Role of β-ARs Signaling in Social Defeat-Induced Reinstatement
Aiming to investigate the involvement of β-ARs signaling in stress-induced reinstatement, a subset of mice received an acute injection of propranolol (10 mg/kg, i.p.) 30 min prior to the social defeat procedure. Control animals received vehicle injections. Fifteen minutes after exposure to psychosocial stress, all animals were subjected to the reinstatement test.
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Understanding whether stress triggers the retrieval of drug-withdrawal memories through activation of the sympathetic nervous system is important for developing novel therapeuti...
- Raw artifact
- Per-sample or per-animal endpoint measurements collected during the experiment
- Processed artifact
- Structured table with cleaned measurements ready for comparison
- Reported as
- Summary statistics and between-group or across-timepoint comparisons
CPA expression was evaluated on day 5 under drug-free conditions by using the same procedure as in the pre-test. The time spent in the naloxone-paired chamber was recorded and r...
- Raw artifact
- Per-sample or per-animal endpoint measurements collected during the experiment
- Processed artifact
- Structured table with cleaned measurements ready for comparison
- Reported as
- Summary statistics and between-group or across-timepoint comparisons
Analysis plan
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Acquisition
Collect raw experimental outputs with enough metadata to preserve sample identity, condition, and timing.
inferred from protocolPreprocessing / cleaning
Extinction sessions began 24 h after the CPA post-test and were conducted daily for 20-25 days until extinction criteria were met.
from paperScoring or quantification
Quantify the primary readouts for this experiment: Understanding whether stress triggers the retrieval of drug-withdrawal memories through activation of the sympathetic nervous system is important for developing novel therapeuti...; CPA expression was evaluated on day 5 under drug-free conditions by using the same procedure as in the pre-test. The time spent in the naloxone-paired chamber was recorded and r....
from paperStatistical comparison
Extinction sessions began 24 h after the CPA post-test and were conducted daily for 20-25 days until extinction criteria were met. Extinction was confirmed using paired t-...
from paperReporting output
Report representative outputs alongside summary comparisons for Understanding whether stress triggers the retrieval of drug-withdrawal memories through activation of the sympathetic nervous system is important for developing novel therapeuti..., CPA expression was evaluated on day 5 under drug-free conditions by using the same procedure as in the pre-test. The time spent in the naloxone-paired chamber was recorded and r....
inferred from protocolStructured statistical methods
Extinction sessions began 24 h after the CPA post-test and were conducted daily for 20-25 days until extinction criteria were met. Extinction was confirmed using paired t-...
source structuredSource and audit
What supports the facts on this page?
Evidence quotes (6)
Understanding whether stress triggers the retrieval of drug-withdrawal memories through activation of the sympathetic nervous system is important for developing novel therapeutic strategies to prevent relapse. Muscle activity measurements offer a potential readout of such stress responses. In this study, a paradigm to measure the re-emergence of an aversive contextual memory induced by physical (restraint and tail-pinch) and psychosocial (social defeat) stressors was applied in mice previously subjected to the CPA procedure. The aim of this research was to expand knowledge about the role of β-ARs in the retrieval of aversive memories and to assess the potential of β-AR antagonism to mitigate physical and/or psychosocial stress-induced negative memories that contribute to possible relapse.
Adult male C57BL/6J mice ( n = 179), which weighed 25-30 g at the beginning of the experiment, were obtained from the Animal Facilities of the University of Murcia (CEIB), Murcia, Spain. These animals were divided into groups of 4-6 per cage under controlled temperature conditions, with access ad libitum to food and water. A 12-h light/dark cycle was established throughout the study. Prior to the experimental procedures, mice were handled and exposed to the testing environment for at least one week. All procedures in this study have adhered to the European Communities Council Directive 2010/63/EU (22 September 2010) and were approved by the Animal Ethics Committee of the University of Murcia (CEEA 534/2019).
Mice were randomly assigned to one of two treatment conditions: chronic saline ( n = 86) or chronic morphine ( n = 93). With the purpose of inducing dependence on morphine, the drug was administered intraperitoneally once daily in escalating doses: 10 mg/kg on day 1, 30 mg/kg on day 2, 50 mg/kg on day 3, and 60 mg/kg on day 4. This dosage has been previously validated to induce opioid tolerance and dependence [,,, ]. Instead, control animals received equivalent volumes of saline following the same schedule. Subsequently, animals were randomly assigned to acute treatment conditions (vehicle or propranolol) and according to stress type (physical, psychosocial, or control group). Behavioral scoring was performed blind to treatment, thereby strengthening the rigor of the experimental design.
Extinction sessions began 24 h after the CPA post-test and were conducted daily for 20-25 days until extinction criteria were met. Extinction was confirmed using paired t-test analyses comparing post-test and final extinction values. The procedure during each session started when the mice were placed in the central passageway and allowed free access to all chambers for 15 min. Time spent in the naloxone-paired chamber and the number of crossings between compartments were recorded.
Reinstatement refers to the re-emergence of a previously extinguished conditioned response following exposure to an unconditioned stimulus-such as a drug, drug-associated cues, or stressors-when extinction has previously occurred [, ]. In the present study, reinstatement of naloxone-induced CPA was evaluated one day after extinction by exposing mice to either an acute social defeat or a physical stressor (restraint or tail-pinch). The reinstatement test followed the same procedure as the pre-test, post-test, and extinction sessions, allowing animals free access to all compartments of the apparatus for 15 min. To minimize contextual associations, all reinstatement procedures were conducted in a room distinct from that used during conditioning and extinction phases, thereby ensuring a non-contingent spatial context relative to the original drug-paired environment.
Aiming to investigate the involvement of β-ARs signaling in stress-induced reinstatement, a subset of mice received an acute injection of propranolol (10 mg/kg, i.p.) 30 min prior to the social defeat procedure. Control animals received vehicle injections. Fifteen minutes after exposure to psychosocial stress, all animals were subjected to the reinstatement test.
Machine-readable layer
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