Exposure to endocrine-disrupting chemicals (EDCs) during gestation influences development of the F1 generation offspring and can result in disease and dysfunction in adulthood. Limited evidence ...suggests consequences on the F2 generation, exposed as germ cells within the F1 fetus. These F2s provide a unique window into the programming effects of EDCs.
This study assessed intergenerational effects of EDC exposure on adult physiology and behavior in Sprague-Dawley rats.
Pregnant rats were exposed to either a polychlorinated biphenyl (PCB) mixture, Aroclor 1,221 (A1221), the fungicide vinclozolin (VIN), or the vehicle (VEH) (6% dimethylsulfoxide in sesame oil) alone. A1221 is weakly estrogenic, while VIN is antiandrogenic, enabling us to compare different classes of EDCs. The F1 male and female offspring were bred to generate the paternal- and maternal-lineage F2 generation. This F2 generation was assessed for physiological outcomes, ultrasonic vocalizations (USVs), and sexual behavior in adulthood.
Each EDC caused phenotypic effects in a sex- and lineage-dependent manner. The most robustly affected group was the paternal-lineage males. F2 VIN paternal male descendants had increased body weight throughout the lifespan, lower concentrations of circulating estradiol, and lower adrenal and testicular indices. Both VIN and A1221 paternal-lineage males also exhibited the greatest number of changes in the characteristics of USVs in response to an opposite-sex animal and changes in sexual behaviors in a mating test.
Exposure of rats to EDCs at the germ cell stage led to differences in the physiological and behavioral phenotype later in life, especially in males. This finding has implications for multigenerational physiological and reproductive health in wildlife and humans. https://doi.org/10.1289/EHP3550.
Celotno besedilo
Dostopno za:
CEKLJ, DOBA, IZUM, KILJ, NUK, OILJ, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK, VSZLJ
•Automated program to analyze ultrasonic vocalization (USV) sound files was developed.•Highly significant correlation between program output and human assessments.•Program completes analyses in ...approximately 0.3% of time compared to manual methods.•USVs shown as useful index for detecting cocaine-associated individual differences.•Statistical analyses reveal cocaine-associated USVs predictive of cocaine intake.
Human emotion is a crucial component of drug abuse and addiction. Ultrasonic vocalizations (USVs) elicited by rodents are a highly translational animal model of emotion in drug abuse studies. A major roadblock to comprehensive use of USV data is the overwhelming burden to attain accurate USV assessment in a timely manner. One of the most accurate methods of analyzing USVs, human auditory detection with simultaneous spectrogram inspection, requires USV sound files to be played back 4% normal speed.
WAAVES (WAV-file Automated Analysis of Vocalizations Environment Specific) is an automated USV assessment program utilizing MATLAB's Signal and Image Processing Toolboxes in conjunction with a series of customized filters to separate USV calls from background noise, and accurately tabulate and categorize USVs as flat or frequency-modulated (FM) calls. In the current report, WAAVES functionality is demonstrated by USV analyses of cocaine self-administration data collected over 10 daily sessions.
WAAVES counts are significantly correlated with human auditory counts (r(48)=0.9925; p<0.001). Statistical analyses used WAAVES output to examine individual differences in USV responses to cocaine, cocaine-associated cues and relationships between USVs, cocaine intake and locomotor activity.
WAAVES output is highly accurate and provides tabulated data in approximately 0.3% of the time required when using human auditory detection methods.
The development of a customized USV analysis program, such as WAAVES streamlines USV assessment and enhances the ability to utilize USVs as a tool to advance drug abuse research and ultimately develop effective treatments.
Short 50-kilohertz (kHz) range frequency-modulated ultrasonic vocalizations (USVs) produced by rats and mice are unconditionally elicited by drugs of abuse or electrical stimulation that increase ...dopamine activity in the nucleus accumbens, and it has been suggested that they reflect “positive affect” or incentive motivational states associated with appetitive behavior. The repeated administration of amphetamine is known to not only produce “psychomotor” sensitization, but also to facilitate a number of appetitive behaviors, including conditioned drug pursuit behavior. We were interested, therefore, in whether amphetamine-induced 50-kHz USVs would also increase with repeated drug exposure. USV recordings were made during 5-min sessions immediately after a saline infusion, and again 4–5
h later after 1.0
mg/kg intravenous amphetamine exposure. These sessions took place every other day over a 5-day period. A challenge dose of 1.0
mg/kg amphetamine was administered 2 weeks later to determine whether sensitization would persist. The initial amphetamine infusion increased 50-kHz USVs relative to the saline infusion. This effect was enhanced over trials and during the amphetamine challenge 2 weeks later. Classification of 50-kHz range call types revealed that complex frequency-modulated trill calls were sensitized by amphetamine, but not flat 50-kHz calls. It is possible that 50-kHz USV recordings could provide a potentially valuable behavioral measure of sensitization linked to enhanced incentive salience and increased tendency to self-administer drugs of abuse.
Ultrasonic vocalization (USV) in the 50-kHz range occurs in rats immediately upon first-time exposure to cocaine or amphetamine, and rapidly increases with repetitive drug exposure at the same dose. ...This sensitized positive-affect response to these drugs of abuse is persistent in that the peak level of USVs again appears when the drug is reintroduced after several weeks of drug discontinuation. The present study explored whether with enough experience USVs might be elicited, and gradually escalate, in anticipation of impending drug delivery. Rats were trained to self-administer (SA) cocaine intravenously by lever pressing 5 days per week for 4 weeks. Yoked rats received experimenter-delivered cocaine matching that of SA rats. USVs and locomotor activity were recorded during each 10-min period prior to 60-min drug access sessions. Extinction trials in which drug access was denied were then carried out over an additional 4-week period. After about a week of cocaine experience, both the SA and yoked groups began to progressively increase USVs when placed in an environment that predicted forthcoming drug exposure. Extinction of anticipatory calls and locomotion occurred over days after drug access ended. USVs may be a useful model for specifically investigating the neural basis of drug anticipation and aid in developing and assessing new addiction treatment strategies for reducing craving and relapse.
Ultrasonic vocalizations (USVs) are well-established markers of motivational and emotional status. Recent work from our lab has provided novel evidence for a role of USVs in models of ethanol (EtOH) ...use. For instance, USV acoustic characteristics can be used to accurately discriminate between rats selectively bred for high EtOH intake (e.g., alcohol-preferring (P) and high-alcohol-drinking (HAD)) versus EtOH-avoiding (e.g., alcohol-non-preferring (NP) and low-alcohol-drinking (LAD)) strains, as well as differentiate between male and female rats. In the present study we sought to explore the effect of age and alcohol availability on spontaneously emitted 50–55 kHz frequency modulated (FM) and 22–28 kHz USVs in adult, male Long–Evans rats. With the hypothesis that age and alcohol experience influence spontaneous USV emissions, we examined USV data collected across a 24-week intermittent EtOH access experiment in male Long–Evans rats. USV counts and acoustic characteristic (i.e., mean frequency, duration, bandwidth and power) data revealed distinct age-dependent phenotypes in both 50–55 kHz FM and 22–28 kHz USV transmission patterns that were modulated by EtOH exposure. These results highlight the influence of age and EtOH experience on the unique emotional phenotypes of male Long–Evans rats.
Heightened emotional states increase impulsive behaviors such as excessive ethanol consumption in humans. Though positive and negative affective states in rodents can be monitored in real-time ...through ultrasonic vocalization (USV) emissions, few animal studies have focused on the role of emotional status as a stimulus for initial ethanol drinking. Our laboratory has recently developed reliable, high-speed analysis techniques to compile USV data during multiple-hour drinking sessions. Since High Alcohol Drinking (HAD-1) rats are selectively bred to voluntarily consume intoxicating levels of alcohol, we hypothesized that USVs emitted by HAD-1 rats would reveal unique emotional phenotypes predictive of alcohol intake and sensitive to alcohol experience. In this study, male HAD-1 rats had access to water, 15% and 30% EtOH or water only (i.e., Controls) during 8 weeks of daily 7-h drinking-in-the-dark (DID) sessions. USVs, associated with both positive (i.e., 50-55 kHz frequency-modulated or FM) and negative (i.e., 22-28 kHz) emotional states, emitted during these daily DID sessions were examined. Findings showed basal 22-28 kHz USVs were emitted by both EtOH-Naïve (Control) and EtOH-experienced rats, alcohol experience enhanced 22-28 kHz USV emissions, and USV acoustic parameters (i.e., mean frequency in kHz) of both positive and negative USVs were significantly suppressed by chronic alcohol experience. These data suggest that negative affective status initiates and maintains excessive alcohol intake in selectively bred HAD-1 rats and support the notion that unprovoked emissions of negative affect-associated USVs (i.e., 22-28 kHz) predict vulnerability to excessive alcohol intake in distinct rodent models.
Rationale
Adult rats often produce 50-kHz ultrasonic vocalizations (USVs), particularly the frequency-modulated varieties, in appetitive situations. These calls are thought by some to reflect ...positive affective states and the reinforcing value of drugs such as amphetamine and cocaine.
Objective
The objective of this study was to determine whether the number of unconditioned 50-kHz USVs elicited by amphetamine predicts the development and/or magnitude of drug-conditioned motivation.
Methods
In three experiments, we recorded USVs before and after injections of 1 mg/kg amphetamine (i.v. or i.p.) administered once per session. Rats were categorized as “high callers” or “low callers” according to individual differences in the number of 50-kHz USVs elicited by their first amphetamine injection. We examined the conditioned appetitive behavior and conditioned place preference (CPP) that emerged in high and low callers after repeated pairings of amphetamine with specific contexts. We also examined whether amphetamine-induced calling was affected by treatment within an unfamiliar (test chamber) versus familiar (home cage) context.
Results
Within an unfamiliar environment, the high callers consistently produced more amphetamine-induced 50-kHz USVs than the low callers. Compared to the low callers, high callers showed significantly greater amphetamine CPP as well as enhanced conditioned 50-kHz USVs and locomotor activity during anticipation of amphetamine. Individual differences were stable when amphetamine was administered in test chambers, but when it was administered in home cages, low callers showed an increase in 50-kHz calling that matched the high callers.
Conclusions
These findings suggest that individual differences in drug-induced USVs can reveal environment-sensitive traits involved in drug-related appetitive motivation.
Rationale
Subtypes of 50-kHz ultrasonic vocalizations (USVs) in rats are thought to reflect positive affect and occur with cocaine or amphetamine delivery. In contexts predicting forthcoming cocaine, ...pre-drug anticipatory USVs are initially minimal during daily sessions but gradually escalate over several weeks, presumably as the animal learns to expect and look forward to impending drug access. To gain more insight into motivational aspects of cocaine intake in animal models of drug dependence studies, it is important to compare experience-dependent changes in lever response rate, USVs, and locomotion during cocaine conditioning and extinction trials.
Objective
To address whether cocaine-induced increases in lever responding and locomotor activity correspond with USV production. The study also determined whether short-term cocaine and context deprivation effects could be detected during conditioning or extinction.
Methods
Rats underwent 20 days of 60-min sessions of self- or yoked administration of cocaine (0.75 mg/kg/infusion, i.v.), followed by 19 days of extinction training (8 weeks total, weekends off).
Results
Lever responding for cocaine and cocaine-induced locomotor activity increased across conditioning sessions. In contrast, the number of frequency modulated 50-kHz USVs evoked in response to cocaine infusion decreased with cocaine experience, suggesting perhaps tolerance to the rewarding properties of the drug. In addition, USVs but not lever pressing or locomotion are affected after brief periods of drug and/or drug context abstinence.
Conclusions
Except for initial drug exposure, increased cocaine seeking during cocaine delivery could reflect either enhanced drug motivation or the development of drug tolerance, but not enhanced positive affect.
Background
Negative emotional status and adverse emotional events increase vulnerability to alcohol abuse. Ultrasonic vocalizations (USVs) emitted by rats are a well‐established model of emotional ...status that can reflect positive or negative affective responses in real time. Most USV studies assess counts, yet each USV is a multidimensional data point characterized by several acoustic characteristics that may provide insight into the neurocircuitry underlying emotional response.
Methods
USVs emitted from selectively bred alcohol‐naïve and alcohol‐experienced alcohol‐preferring and nonpreferring rats (P and NP rats) were recorded during 4‐hour sessions on alternating days over 4 weeks. Linear mixed modeling (LMM) and linear discriminant analysis (LDA) were applied to USV acoustic characteristics (e.g., frequency, duration, power, and bandwidth) of negative affect (22 to 28 kilohertz kHz)‐ and positive (50 to 55 kHz) affect‐related USVs.
Results
Hundred percent separation between alcohol‐naïve P and NP rats was achieved through a linear combination (produced by LDA) of USV acoustic characteristics of 22‐ to 28‐kHz USVs, whereas poor separation (36.5%) was observed for 50‐ to 55‐kHz USVs. 22‐ to 28‐kHz LDA separation was high (87%) between alcohol‐experienced P and NP rats, but was poor for 50‐ to 55‐kHz USVs (57.3%). USV mean frequency and duration were the highest weighted characteristics in both the naïve and experienced 22‐ to 28‐kHz LDA representations suggesting that alcohol experience does not alter the representations. LMM analyses of 22‐ to 28‐kHz USV acoustic characteristics matched the LDA results. Poor LDA separation was observed between alcohol‐naïve and alcohol‐experienced P rats for both 22‐ to 28‐kHz and 50‐ to 55‐kHz USVs.
Conclusions
Advanced statistical analysis of negative affect‐associated USV data predicts future behaviors of excessive alcohol drinking and alcohol avoidance in selectively bred rats. USV characteristics across rat lines reveal affect‐related motivation to consume alcohol and may predict neural pathways mediating emotional response. Further characterization of these differences could delineate particular neurocircuitry and methods to ameliorate dysregulated emotional states often observed in human alcohol abusers.
Diagram depicts a mixed population of rats categorized into P and NP groups based solely on 22 kHz USV record data. The procedure includes USV recordings, analyses of the vocalization recordings using the WAAVES program (Reno et al. 2013) and tabulation of 22–28 kHz USV acoustic characteristics (mean frequency, duration, bandwidth and power). Categorization of P versus NP groups is based solely on a linear combination of acoustic characteristics.
Excessive alcohol consumption has a vast, negative impact on society. Rodent models have been successful in furthering our understanding of the biological underpinnings that drive alcohol ...consumption. Rodents emit ultrasonic vocalizations (USVs) that are each composed of several acoustic characteristics (e.g., frequency, duration, bandwidth, power). USVs reflect neurotransmitter activity in the ascending limb of the mesolimbic dopaminergic and cholinergic neurotransmitter systems and serve as noninvasive, real-time biomarkers of dopaminergic and cholinergic neurotransmission in the limbic system. In the present study, we recorded spontaneously emitted USVs from alcohol-naïve Long-Evans (LE) rats and then measured their alcohol intake. We compared the USV acoustic characteristics and alcohol consumption data from these LE rats with previously published data from selectively bred high-alcohol (P and HAD-1) and low-alcohol (NP and LAD-1) drinking lines from studies with the same experimental method. Predictive analytic techniques were applied simultaneously to this combined data set and revealed that (a) USVs emitted by alcohol-naïve rats accurately discriminated among high-alcohol consuming, LE, and low-alcohol consuming rat lines, and (b) future alcohol consumption in these same rat lines was reliably predicted from the USV data collected in an alcohol-naïve state. To our knowledge, this is the first study to show that alcohol consumption is predicted directly from USV profiles of alcohol-naïve rats. Because USV acoustic characteristics are sensitive to underlying neural activity, these findings suggest that baseline differences in mesolimbic cholinergic and dopaminergic tone could determine the propensity for future alcohol consumption in rodents.
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