Behaviorally conditioned effects of psychoactive drugs in experimental animals: What we have learned from nearly a century of research and what remains to be learned.

Continuous treatment with drugs is a crucial requirement for managing various clinical conditions, including chronic pain and neuropsychiatric disorders such as depression or schizophrenia. Associative learning processes, i.e. Pavlovian conditioning, can play an important role for the effects of drugs and could open new avenues for optimizing patient treatment. In this narrative literature review, we summarize available data in experimental animals regarding the behaviorally conditioned effects of psychostimulants such as d-amphetamine and cocaine, the dopamine receptor agonist apomorphine, the dopamine receptor antagonist haloperidol, morphine and antidepressant drugs. In each section, the drug under discussion is briefly introduced, followed by a detailed examination of conditioning features, including doses and dosing regimens, characteristics of the conditioning process such as test environments or specific conditioned stimuli, testing and conditioned response characteristics, possible extinction or reconditioning or reversal training, neural mechanisms, and finally, the potential clinical relevance of the research area related to the drug. We focus on key outcomes, delve into methodical issues, identify gaps in current knowledge, and suggest future research directions.

Forty-kHz ultrasonic vocalizations of rat pups predict adult behavior in the elevated plus-maze behavior but not the effect of cocaine on 50-kHz ultrasonic vocalizations.

Ultrasonic vocalizations (USV) are emitted by both young pups and adult rats to convey positive or negative emotional states. These USV manifestations are contingent on factors including developmental stage, situational requirements, and individual dispositions. Pups emit 40-kHz USV when separated from their mother and litter, which function to elicit maternal care. Conversely, adult rats can produce 50-kHz USV in response to stimuli that elicit reward-related states, including natural rewards, stimulant drugs, and reward-predictive stimuli. The present study aims to investigate whether pup 40-kHz USV can serve as predictors of behaviors related to positive or negative states in adult rats. Both male and female Wistar pups were initially tested on the 11th postnatal day and subsequently in adulthood. There was no significant difference in the number of 40-kHz ultrasonic vocalizations between male and female pups. However, cocaine elicited more 50-kHz USV and hyperactivity in adult females compared to males. Notably, cocaine increased the proportion of step and trill USV subtypes in both adult males and females. Interestingly, this effect of cocaine was stronger in females that were in the diestrus, compared to the estrus phase. In males, a significant positive correlation was found between pup 40-kHz USV and lower anxiety scores in adult male but not female rats tested on the elevated plus-maze test. Furthermore, no significant correlation was found between pup 40-kHz and adult 50-kHz USV in both males and females, whether in undrugged (saline) or in cocaine-treated rats. It is possible that the 40-kHz USV emitted by pups predicted reduced anxiety-like behavior only for male rats because they could elicit maternal care directed specifically to male pups. These findings suggest that 40-kHz USV can serve as an indicator of the emotional link between the rat mother and male pups. Indeed, this suggests that maternal care exerts a positive influence on the emotional state during adulthood.

Biperiden reverses the increase in 50-kHz ultrasonic vocalizations but not the increase in locomotor activity induced by cocaine.

Cocaine use disorder (CUD) is a worldwide public health problem, associated with severe psychosocial and economic impacts. Currently, no FDA-approved treatment is available for CUD. However, an emerging body of evidence from clinical and preclinical studies suggests that biperiden, an M1 muscarinic receptor antagonist, presents potential therapeutic use for CUD. These studies have suggested that biperiden may reduce the reinforcing effects of cocaine. It is well established that rodents emit 50-kHz ultrasonic vocalizations (USV) in response to natural rewards and stimulant drugs, including cocaine. Nonetheless, the effects of biperiden on the cocaine-induced increase of 50-kHz USV remains unknown. Here, we hypothesized that biperiden could antagonize the acute effects of cocaine administration on rat 50-kHz USV. To test this hypothesis, adult male Wistar rats were divided into four experimental groups: saline, 5 mg/kg biperiden, 10 mg/kg cocaine, and biperiden/cocaine (5 and 10 mg/kg, i.p., respectively). USV and locomotor activity were recorded in baseline and test sessions. As expected, cocaine administration significantly increased the number of 50-kHz USV. Biperiden administration effectively antagonized the increase in 50-kHz USV induced by cocaine. Cocaine administration also increased the emission of trill and mixed 50 kHz USV subtypes and this effect was antagonized by biperiden. Additionally, we showed that biperiden did not affect the cocaine-induced increase in locomotor activity, although biperiden administration per se increased locomotor activity. In conclusion, our findings indicate that administering biperiden acutely reduces the positive affective effects of cocaine, as demonstrated by its ability to inhibit the increase in 50-kHz USV.

Acute anxiogenic effects of escitalopram are associated with mild alterations in D-amphetamine-induced behavior and social approach evoked by playback of 50-kHz ultrasonic vocalizations in rats.

Rats communicate through auditory signals in the ultrasonic range, so-called ultrasonic vocalizations (USV). Short, high-frequency 50-kHz USV are associated with positive affective states and are emitted in appetitive situations, often rewarding social interactions, such as rough-and-tumble play and mating. Exaggerated levels of 50-kHz USV emission can be observed in response to psychostimulants, most notably d-amphetamine (AMPH). There is robust evidence suggesting that 50-kHz USV serve as affiliative signals and help to maintain or re-establish social proximity. A key neurotransmitter involved in behavioral regulation is serotonin (5-hydroxytryptamine, 5-HT). This includes both, the regulation of anxiety-related behavior and ultrasonic communication. Here, we show that acute treatment with the selective 5-HT reuptake inhibitor (SSRI) escitalopram (ESC) leads to increased anxiety-related behavior in the elevated plus maze and tested whether such acute anxiogenic effects of ESC result in alterations in ultrasonic communication in sender and/or receiver. To this aim, we conducted a dose-response study in male rats and assessed AMPH-induced hyperactivity and 50-kHz ultrasonic calling in the sender and social approach behavior evoked by playback of pro-social 50-kHz USV in the receiver. Acute ESC treatment affected both, sender and receiver. This was reflected in a lack of AMPH-induced changes in acoustic features of 50-kHz USV and absence of social exploratory behavior evoked by 50-kHz USV playback, respectively. Albeit the SSRI effects were relatively mild, this supports the notion that the 5-HT system is involved in the regulation of a key aspect of the social behavior repertoire of rodents, namely socio-affective communication through 50-kHz USV.

Wildtype peers rescue social play and 50-kHz ultrasonic vocalization deficits in juvenile female Cacna1c heterozygous rats.

Background: Healthy brain development depends on early social practices and experiences. The risk gene CACNA1C is implicated in numerous neuropsychiatric disorders, in which key characteristics include deficits in social functioning and communication. Recently, we reported sex-dependent impairments in social behavior and ultrasonic vocalizations (USV) in juvenile heterozygous Cacna1c+/- (HET) rats. Specifically, HET females displayed increases in rough-and-tumble play that eliminated the typically observed sex difference between male and female rats. Interestingly, female wild-type Cacna1c+/+ (WT) pairs also showed a similar increase in social play when housed with HET females, suggesting their behavior may be influenced by HET cage mates. This indicates that the genetic makeup of the social environment related to Cacna1c can influence social play, yet systematic studies are lacking. Methods: In the present study, we housed juvenile females in MIXED- or SAME-genotype cages and tested them in a social play paradigm with a same- and opposite-genotype partner. Results: The results show that the early social environment and the genotype of the play partner influence social play and 50-kHz USV emission. Experience with a WT play partner appears necessary for HET females to show comparable levels of play and 50-kHz USV emission. Same-genotype HET pairs played less and emitted fewer 50-kHz USV than same-genotype WT or opposite-genotype pairs; however, we found that the decrease in social play and 50-kHz USV in HET pairs can be rescued by playing with a WT partner. The effect was particularly prominent when the first play partner was WT, as we found it increased play and 50-kHz USV emission in all subsequent interactions with ensuing partners. Conclusion: These findings suggest that the genetic makeup related to the social environment and/or social peers influences social play in Cacna1c+/- haploinsufficient rats. Specifically, our results show that WT peers can rescue behavior and communication alterations in Cacna1c female rats. Our findings have important implications because they show that the genetic makeup of the social environment can divulge phenotypic changes in genetic rat models of neuropsychiatric disorders.

Behavioral analysis in laboratory rats: Challenges and usefulness of 50-kHz ultrasonic vocalizations.

Many rodent species emit and detect vocalizations in the ultrasonic range. Rats use three classes of ultrasonic vocalizations depending on developmental stage, experience and the behavioral situation. Calls from one class emitted by juvenile and adult rats, the so-called 50-kHz calls, are typical for appetitive and social situations. This review provides a brief historical account on the introduction of 50-kHz calls in behavioral research followed by a survey of their scientific applications focusing on the last five years, where 50-kHz publications reached a climax. Then, specific methodological challenges will be addressed, like how to measure and report 50-kHz USV, the problem of assignment of acoustic signals to a specific sender in a social situation, and individual variability in call propensity. Finally, the intricacy of interpreting 50-kHz results will be discussed focusing on the most prevalent ones, namely as communicative signals and/or readouts of the sender's emotional status.

Calcium Handling Remodeling Underlies Impaired Sympathetic Stress Response in Ventricular Myocardium from Cacna1c Haploinsufficient Rats.

CACNA1C encodes the pore-forming α1C subunit of the L-type Ca2+ channel, Cav1.2. Mutations and polymorphisms of the gene are associated with neuropsychiatric and cardiac disease. Haploinsufficient Cacna1c+/- rats represent a recently developed model with a behavioral phenotype, but its cardiac phenotype is unknown. Here, we unraveled the cardiac phenotype of Cacna1c+/- rats with a main focus on cellular Ca2+ handling mechanisms. Under basal conditions, isolated ventricular Cacna1c+/- myocytes exhibited unaltered L-type Ca2+ current, Ca2+ transients (CaTs), sarcoplasmic reticulum (SR) Ca2+ load, fractional release, and sarcomere shortenings. However, immunoblotting of left ventricular (LV) tissue revealed reduced expression of Cav1.2, increased expression of SERCA2a and NCX, and augmented phosphorylation of RyR2 (at S2808) in Cacna1c+/- rats. The ß-adrenergic agonist isoprenaline increased amplitude and accelerated decay of CaTs and sarcomere shortenings in both Cacna1c+/- and WT myocytes. However, the isoprenaline effect on CaT amplitude and fractional shortening (but not CaT decay) was impaired in Cacna1c+/- myocytes exhibiting both reduced potency and efficacy. Moreover, sarcolemmal Ca2+ influx and fractional SR Ca2+ release after treatment with isoprenaline were smaller in Cacna1c+/- than in WT myocytes. In Langendorff-perfused hearts, the isoprenaline-induced increase in RyR2 phosphorylation at S2808 and S2814 was attenuated in Cacna1c+/- compared to WT hearts. Despite unaltered CaTs and sarcomere shortenings, Cacna1c+/- myocytes display remodeling of Ca2+ handling proteins under basal conditions. Mimicking sympathetic stress with isoprenaline unmasks an impaired ability to stimulate Ca2+ influx, SR Ca2+ release, and CaTs caused, in part, by reduced phosphorylation reserve of RyR2 in Cacna1c+/- cardiomyocytes.

Paradoxical kinesia may no longer be a paradox waiting for 100 years to be unraveled.

Parkinson's disease (PD) is a progressive neurodegenerative disorder mainly characterized by bradykinesia and akinesia. Interestingly, these motor disabilities can depend on the patient emotional state. Disabled PD patients remain able to produce normal motor responses in the context of urgent or externally driven situations or even when exposed to appetitive cues such as music. To describe this phenomenon Souques coined the term "paradoxical kinesia" a century ago. Since then, the mechanisms underlying paradoxical kinesia are still unknown due to a paucity of valid animal models that replicate this phenomenon. To overcome this limitation, we established two animal models of paradoxical kinesia. Using these models, we investigated the neural mechanisms of paradoxical kinesia, with the results pointing to the inferior colliculus (IC) as a key structure. Intracollicular electrical deep brain stimulation, glutamatergic and GABAergic mechanisms may be involved in the elaboration of paradoxical kinesia. Since paradoxical kinesia might work by activation of some alternative pathway bypassing basal ganglia, we suggest the IC as a candidate to be part of this pathway.

Using expectation violation models to improve the outcome of psychological treatments.

Expectations are a central maintaining mechanism in mental disorders and most psychological treatments aim to directly or indirectly modify clinically relevant expectations. Therefore, it is crucial to examine why patients with mental disorders maintain dysfunctional expectations, even in light of disconfirming evidence, and how expectation-violating situations should be created in treatment settings to optimize treatment outcome and reduce the risk of treatment failures. The different psychological subdisciplines offer various approaches for understanding the underlying mechanisms of expectation development, persistence, and change. Here, we convey recommendations on how to improve psychological treatments by considering these different perspectives. Based on our expectation violation model, we argue that the outcome of expectation violation depends on several characteristics: features of the expectation-violating situation; the dynamics between the magnitude of expectation violation and cognitive immunization processes; dealing with uncertainties during and after expectation change; controlled and automatic attention processes; and the costs of expectation changes. Personality factors further add to predict outcomes and may offer a basis for personalized treatment planning. We conclude with a list of recommendations derived from basic psychology that could contribute to improved treatment outcome and to reduced risks of treatment failures.

Contingent Social Interaction Does Not Prevent Habituation towards Playback of Pro-Social 50-kHz Calls: Behavioral Responses and Brain Activation Patterns.

Rats, which are highly social animals, are known to communicate using ultrasonic vocalizations (USV) in different frequency ranges. Calls around 50 kHz are related to positive affective states and promote social interactions. Our previous work has shown that the playback of natural 50-kHz USV leads to a strong social approach response toward the sound source, which is related to activation in the nucleus accumbens. In male Wistar rats, the behavioral response habituates, that is, becomes weaker or is even absent, when such playback is repeated several days later, an outcome found to be memory-dependent. Here, we asked whether such habituation is due to the lack of a contingent social consequence after playback in the initial test and whether activation of the nucleus accumbens, as measured by c-fos immunohistochemistry, can still be observed in a retest. To this end, groups of young male Wistar rats underwent an initial 50-kHz USV playback test, immediately after which they were either (1) kept temporarily alone, (2) exposed to a same-sex juvenile, or (3) to their own housing group. One week later, they underwent a retest with playback; this time not followed by social consequences but by brain removal for c-fos immunohistochemistry. Consistent with previous reports, behavioral changes evoked by the initial exposure to 50-kHz USV playback included a strong approach response. In the retest, no such response was found, irrespective of whether rats had experienced a contingent social consequence after the initial test or not. At the neural level, no substantial c-fos activation was found in the nucleus accumbens, but unexpected strong activation was detected in the anterior cingulate cortex, with some of it in GABAergic cells. The c-fos patterns did not differ between groups but cell numbers were individually correlated with behavior, i.e., rats that still approached in response to playback in the retest showed more activation. Together, these data do not provide substantial evidence that the lack of a contingent social consequence after 50-kHz USV playback accounts for approach habituation in the retest. Additionally, there is apparently no substantial activation of the nucleus accumbens in the retest, whereas the exploratory findings in the anterior cingulate cortex indicate that this brain area might be involved when individual rats still approach 50-kHz USV playback.

Bipolar-associated miR-499-5p controls neuroplasticity by downregulating the Cav1.2 subunit CACNB2.

Bipolar disorder (BD) is a chronic mood disorder characterized by manic and depressive episodes. Dysregulation of neuroplasticity and calcium homeostasis are frequently observed in BD patients, but the underlying molecular mechanisms are largely unknown. Here, we show that miR-499-5p regulates dendritogenesis and cognitive function by downregulating the BD risk gene CACNB2. miR-499-5p expression is increased in peripheral blood of BD patients, as well as in the hippocampus of rats which underwent juvenile social isolation. In rat hippocampal neurons, miR-499-5p impairs dendritogenesis and reduces surface expression and activity of the L-type calcium channel Cav1.2. We further identified CACNB2, which encodes a regulatory ß-subunit of Cav1.2, as a direct functional target of miR-499-5p in neurons. miR-499-5p overexpression in the hippocampus in vivo induces short-term memory impairments selectively in rats haploinsufficient for the Cav1.2 pore forming subunit Cacna1c. In humans, miR-499-5p expression is negatively associated with gray matter volumes of the left superior temporal gyrus, a region implicated in auditory and emotional processing. We propose that stress-induced miR-499-5p overexpression contributes to dendritic impairments, deregulated calcium homeostasis, and neurocognitive dysfunction in BD.

Trigeminal neuropathic pain reduces 50-kHz ultrasonic vocalizations in rats, which are restored by analgesic drugs.

Trigeminal neuralgia (TN) is a severe form of neuropathic pain frequently associated with anxiety. The chronic constriction injury of the infraorbital nerve (CCI-ION) of rodents is a well-established model to study sensory alterations related to TN. However, few studies have addressed the emotional component of pain, which is fundamental to increase its translational capability. Emission of ultrasonic vocalization (USV) is considered a reliable measure of the emotional state of rats. Rats emit 50-kHz USVs in social and appetitive situations, whereas 22-kHz USVs may index a negative state. Studies suggest that persistent pain causes reduction in 50-kHz calls, but this may also indicate anxiety-like behavior. Thus, we hypothesize that CCI-ION would decrease 50-kHz calls and that pharmacological pain relief would restore USVs, without interfering with anxiety-like behavior. On day 15 after surgery, male rats were treated with local lidocaine, midazolam or carbamazepine to determine their effect on facial mechanical hyperalgesia, USV and anxiety-like behavior. The results showed that CCI-ION induced hyperalgesia, which was attenuated by lidocaine or carbamazepine, developed anxiety-like behavior, which was reduced only by midazolam, and displayed a reduced number of 50-kHz calls, compared to sham. Lidocaine and carbamazepine increased 50-kHz calls emitted by CCI-ION rats, but midazolam failed to change them. These data add information on the translational aspects of CCI-ION model and carbamazepine treatment for trigeminal neuropathic pain. Furthermore, they suggest that the reduction of USV in persistent pain conditions is related to spontaneous pain and reinforce the idea that it reflects the emotional component of pain.

Appetitive 50 kHz calls in a pavlovian conditioned approach task in Cacna1c haploinsufficient rats.

We have previously shown that rats emit high-frequency 50 kHz ultrasonic vocalizations (USV) during sign- and goal-tracking in a common Pavlovian conditioned approach task. Such 50 kHz calls are probably related to positive affect and are associated with meso-limbic dopamine function. In humans, the CACNA1C gene, encoding for the α1C subunit of the L-type voltage-gated calcium channel CaV1.2, is implicated in several mental disorders, including mood disorders associated with altered dopamine signaling. In the present study, we investigated sign- and goal-tracking behavior and the emission of 50 kHz USV in Cacna1c haploinsufficent rats in a task where food pellet delivery is signaled by an appearance of an otherwise inoperable lever. Over the course of this Pavlovian training, these rats not only increased their approach to the reward site, but also their rates of pressing the inoperable lever. During subsequent extinction tests, where reward delivery was omitted, extinction patterns differed between reward site (i.e. magazine entries) and lever, since magazine entries quickly declined whereas behavior towards the lever transiently increased. Based on established criteria to define sign- or goal-tracking individuals, no CACNA1C rat met a sign-tracking criterion, since around 42% of rats tested where goal-trackers and the other 58% fell into an intermediate range. Regarding USV, we found that the CACNA1C rats emitted 50 kHz calls with a clear subject-dependent pattern; also, most of them were of a flat subtype and occurred mainly during initial habituation phases without cues or rewards. Compared, to previously published wildtype controls, Cacna1c haploinsufficent rats displayed reduced numbers of appetitive 50 kHz calls. Moreover, similar to wildtype littermate controls, 50 kHz call emission in Cacna1c haploinsufficent rats was intra-individually stable over training days and was negatively associated with goal-tracking. Together, these findings provide evidence in support of 50 kHz calls as trait marker. The finding that Cacna1c haploinsufficent rats show reductions of 50 kHz calls accompanied with more goal-tracking, is consistent with the assumption of altered dopamine signaling in these rats, a finding which supports their applicability in models of mental disorders.

Diazepam attenuates the effects of cocaine on locomotion, 50-kHz ultrasonic vocalizations and phasic dopamine in the nucleus accumbens of rats.

BACKGROUND AND PURPOSE: Currently, there is no effective drug to treat cocaine-use disorder, which affects millions of people worldwide. Benzodiazepines are potential therapeutic candidates, as microdialysis and voltammetry studies have shown that they can decrease dopamine concentrations in the nucleus accumbens of rodents and block the increase in dopamine levels and appetitive 50-kHz ultrasonic vocalizations (USVs) induced by amphetamine in rats. EXPERIMENTAL APPROACH: Here, we tested whether administration of 2.5-mg·kg-1 diazepam (i.p.) in adult male rats could block the effects of 20-mg·kg-1 cocaine (i.p.) on electrically evoked phasic dopamine signals in the nucleus accumbens measured by fast-scan cyclic voltammetry, as well as 50-kHz USV and locomotor activity. KEY RESULTS: Cocaine injection increased evoked dopamine signals up to threefold within 5 min, and the increase was significantly higher than baseline for at least 75 min. The injection of diazepam, 5 min after cocaine, attenuated the cocaine effect by nearly 50%, and this attenuation was maintained for at least 40 min. Behaviourally, cocaine increased the number of appetitive 50-kHz calls by about 12-fold. Diazepam significantly blocked this effect for the entire duration of the session. Also, cocaine-treated rats were more active than controls and diazepam significantly attenuated cocaine-induced locomotion, by up to 50%. CONCLUSION AND IMPLICATIONS: These results suggest that the neurochemical and psychostimulant effects of cocaine can be mitigated by diazepam. LINKED ARTICLES: This article is part of a themed issue on Building Bridges in Neuropharmacology. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v179.8/issuetoc.

Fear Extinction and Predictive Trait-Like Inter-Individual Differences in Rats Lacking the Serotonin Transporter.

Anxiety disorders are associated with a failure to sufficiently extinguish fear memories. The serotonergic system (5-hydroxytryptamine, 5-HT) with the 5-HT transporter (5-HTT, SERT) is strongly implicated in the regulation of anxiety and fear. In the present study, we examined the effects of SERT deficiency on fear extinction in a differential fear conditioning paradigm in male and female rats. Fear-related behavior displayed during acquisition, extinction, and recovery, was measured through quantification of immobility and alarm 22-kHz ultrasonic vocalizations (USV). Trait-like inter-individual differences in novelty-seeking, anxiety-related behavior, habituation learning, cognitive performance, and pain sensitivity were examined for their predictive value in forecasting fear extinction. Our results show that SERT deficiency strongly affected the emission of 22-kHz USV during differential fear conditioning. During acquisition, extinction, and recovery, SERT deficiency consistently led to a reduction in 22-kHz USV emission. While SERT deficiency did not affect immobility during acquisition, genotype differences started to emerge during extinction, and during recovery rats lacking SERT showed higher levels of immobility than wildtype littermate controls. Recovery was reflected in increased levels of immobility but not 22-kHz USV emission. Prominent sex differences were evident. Among several measures for trait-like inter-individual differences, anxiety-related behavior had the best predictive quality.

Social Behavior and Ultrasonic Vocalizations in a Genetic Rat Model Haploinsufficient for the Cross-Disorder Risk Gene Cacna1c.

The top-ranked cross-disorder risk gene CACNA1C is strongly associated with multiple neuropsychiatric dysfunctions. In a recent series of studies, we applied a genomically informed approach and contributed extensively to the behavioral characterization of a genetic rat model haploinsufficient for the cross-disorder risk gene Cacna1c. Because deficits in processing social signals are associated with reduced social functioning as commonly seen in neuropsychiatric disorders, we focused on socio-affective communication through 22-kHz and 50-kHz ultrasonic vocalizations (USV). Specifically, we applied a reciprocal approach for studying socio-affective communication in sender and receiver by including rough-and-tumble play and playback of 22-kHz and 50-kHz USV. Here, we review the findings obtained in this recent series of studies and link them to the key features of 50-kHz USV emission during rough-and-tumble play and social approach behavior evoked by playback of 22-kHz and 50-kHz USV. We conclude that Cacna1c haploinsufficiency in rats leads to robust deficits in socio-affective communication through 22-kHz and 50-kHz USV and associated alterations in social behavior, such as rough-and-tumble play behavior.

Andrographolide blocks 50-kHz ultrasonic vocalizations, hyperlocomotion and oxidative stress in an animal model of mania.

In rats, lisdexamfetamine (LDX) induces manic-like behaviors such as hyperlocomotion and increases in appetitive 50-kHz ultrasonic vocalizations (USV), which are prevented by antimanic drugs, such as lithium. Inhibition of glycogen synthase kinase 3 beta (GSK3ß) and antioxidant activity have been associated with antimanic effects. Thus, the aim of the present study was to evaluate the possible antimanic-like effects of andrographolide (ANDRO), a GSK3ß inhibitor, on LDX-induced hyperlocomotion and 50-kHz USV increases. In addition, the effect of ANDRO was studied on LDX-induced oxidative stress. Lithium was used as positive control. Adult Wistar rats were treated with vehicle, lithium (100 mg/kg i.p., daily) or ANDRO (2 mg/kg i.p., 3 times a week) for 21 days. On the test day, either 10 mg/kg LDX or saline was administered i.p. and USV and locomotor activity were recorded. LDX administration increased the number of 50-kHz calls, as well as locomotor activity. Repeated treatment with lithium or ANDRO prevented these effects of LDX on 50-kHz USV and locomotor activity. LDX increased lipid peroxidation (LPO) levels in rat striatum and both lithium and ANDRO prevented this effect. LPO levels in rat striatum were positively correlated with increases in 50-kHz USV emission as well as hyperlocomotion. In conclusion, the present results indicate that ANDRO has antimanic-like effects, which may be mediated by its antioxidant properties.

Limited generalizability, pharmacological modulation, and state-dependency of habituation towards pro-social 50-kHz calls in rats.

Communication constitutes a fundamental component of mammalian social behavior. Rats are highly social animals and emit 50-kHz ultrasonic vocalizations (USV), which function as social contact calls. Playback of 50-kHz USV leads to strong and immediate social approach responses in receiver rats, but this response is weak or even absent during repeated 50-kHz USV playback. Given the important role of 50-kHz USV in initiating social contact and coordinating social interactions, the occurrence of habituation is highly unexpected. It is not clear why a social signal characterized by significant incentive salience loses its power to change the behavior of the receiver so rapidly. Here, we show that the habituation phenomenon displayed by rats in response to repeated playback of 50-kHz USV (1) is characterized by limited generalizability because it is present in Wistar but not Sprague-Dawley rats, (2) can be overcome by amphetamine treatment, and (3) depends on the subject's internal state.

Sex differences in the acoustic features of social play-induced 50-kHz ultrasonic vocalizations: A detailed spectrographic analysis in wild-type Sprague-Dawley and Cacna1c haploinsufficient rats.

Sexual dimorphisms are widespread in the animal kingdom. At the behavioral level, there is evidence for sex differences in social play behavior. In rats, males typically engage more in rough-and-tumble play than females. One prominent component of the rough-and-tumble play repertoire in rats is the emission of 50-kHz ultrasonic vocalizations (USV). Such 50-kHz USV reflect the rewarding nature of play and serve as socioaffective signals. Here, we provide evidence for sexual dimorphisms within rough-and-tumble play-induced 50-kHz USV in juvenile rats. Specifically, females displayed reduced 50-kHz USV emission during playful interactions. This reduction was associated with changes in 50-kHz USV emission rates and subtype profiles during specific rough-and-tumble components, i.e., pinning, wrestling, and chasing, as well as differences in acoustic parameters. Interestingly, sex differences were modulated by Cacna1c, a gene strongly implicated in major neuropsychiatric disorders, often characterized by prominent sex biases, most notably autism. Specifically, Cacna1c haploinsufficiency affected the emission of 50-kHz USV during rough-and-tumble play in female rats and we provide evidence supporting the notion that such effects of Cacna1c haploinsufficiency are driven by male-typical features of 50-kHz USV emission. This is in line with the hypermasculinized social play repertoire previously observed in juvenile Cacna1c haploinsufficient females.

Response Calls Evoked by Playback of Natural 50-kHz Ultrasonic Vocalizations in Rats.

Rats are highly social animals known to communicate with ultrasonic vocalizations (USV) of different frequencies. Calls around 50 kHz are thought to represent a positive affective state, whereas calls around 22 kHz are believed to serve as alarm or distress calls. During playback of natural 50-kHz USV, rats show a reliable and strong social approach response toward the sound source. While this response has been studied in great detail in numerous publications, little is known about the emission of USV in response to natural 50-kHz USV playback. To close this gap, we capitalized on three data sets previously obtained and analyzed USV evoked by natural 50-kHz USV playback in male juvenile rats. We compared different rat stocks, namely Wistar (WI) and Sprague-Dawley (SD) and investigated the pharmacological treatment with the dopaminergic D2 receptor antagonist haloperidol. These response calls were found to vary broadly inter-individually in numbers, mean peak frequencies, durations and frequency modulations. Despite the large variability, the results showed no major differences between experimental conditions regarding call likelihood or call parameters, representing a robust phenomenon. However, most response calls had clearly lower frequencies and were longer than typical 50-kHz calls, i.e., around 30 kHz and lasting generally around 0.3 s. These calls resemble aversive 22-kHz USV of adult rats but were of higher frequencies and shorter durations. Moreover, blockade of dopamine D2 receptors did not substantially affect the emission of response calls suggesting that they are not dependent on the D2 receptor function. Taken together, this study provides a detailed analysis of response calls toward playback of 50-kHz USV in juvenile WI and SD rats. This includes calls representing 50-kHz USV, but mostly calls with lower frequencies that are not clearly categorizable within the so far known two main groups of USV in adult rats. We discuss the possible functions of these response calls addressing their communicative functions like contact or appeasing calls, and whether they may reflect a state of frustration. In future studies, response calls might also serve as a new read-out in rat models for neuropsychiatric disorders, where acoustic communication is impaired, such as autism spectrum disorder.