Alcohol‐Preferring P Rats Emit Spontaneous 22‐28 kHz Ultrasonic Vocalizations that are Altered by Acute and Chronic Alcohol Experience

Abstract

Background Emotional states are often thought to drive excessive alcohol intake and influence the development of alcohol use disorders (AUDs). To gain insight into affective properties associated with excessive alcohol intake, we utilized ultrasonic vocalization (USV) detection and analyses to characterize the emotional phenotype of selectively bred alcohol-preferring “P rats”; an established animal model of excessive alcohol intake. USVs emitted by rodents have been convincingly associated with positive (50–55 kHz) and negative (22–28 kHz) affective states. Therefore, we hypothesized that 50–55 and 22–28 kHz USV emission patterns in alcohol-preferring P rats would reveal a unique emotional phenotype sensitive to alcohol experience. Methods 50–55 kHz frequency-modulated (FM) and 22–28 kHz USVs elicited from male alcohol-preferring P rats were assessed during access to water, 15% and 30% EtOH (v/v). (EtOH; n=12) or water only (Control; n=4) across 8 weeks of daily drinking-in-the-dark (DID) sessions. Results Spontaneous 22–28 kHz USVs are emitted by alcohol-naïve P rats and are enhanced by alcohol experience. During DID sessions when alcohol was not available (e.g., “EtOH OFF” intervals), significantly more 22–28 kHz than 50–55 kHz USVs were elicited, while significantly more 50–55 kHz than 22–28 kHz USVs were emitted when alcohol was available (e.g., “EtOH ON” intervals). In addition, USV acoustic property analyses revealed chronic effects of alcohol experience on 22–28 kHz USV mean frequency, indicative of lasting alcohol-mediated alterations to neural substrates underlying emotional response. Conclusions Our findings demonstrate that acute and chronic effects of alcohol exposure are reflected in changes in 22–28 and 50–55 kHz USV counts and acoustic patterns. These data support the notion that initiation and maintenance of alcohol intake in P rats may be due to a unique, alcohol-responsive emotional phenotype and further suggest that spontaneous 22–28 kHz USVs serve as behavioral markers for excessive drinking vulnerability.

Alcohol enhances unprovoked 22–28kHz USVs and suppresses USV mean frequency in High Alcohol Drinking (HAD-1) male rats

“…Though this finding is consistent with our recent report on the selectively bred alcohol-preferring P rat [24], spontaneous 22–28 kHz USV emissions have not been reported in any other rat lines. In addition, the mean frequency of positive affect USVs (approx.…”

“…We recently reported ethanol consumption in alcohol-preferring P rats that was lower than previous reports [24]. We attributed lower ethanol consumption levels to the extended pre-experimental handling procedures (e.g., 4 weeks of daily handling sessions) used to decrease anxiogenic and/or negative emotional responses to human touch since other DID studies reporting higher ethanol intake levels do not report a handling phase in their experiments [31–34].…”

“…Recently, our laboratory has recorded USVs from one of these models, the alcohol-preferring (P) rat. During a Drinking in the Dark (DID) experiment, we observed spontaneous 22–28 kHz USV emissions from both alcohol-naïve and alcohol-experienced P rats [24]. In the alcohol literature, 22–28 kHz USVs have been intentionally provoked by exposure to aversive air-puff stimuli in alcohol-dependent animals undergoing alcohol withdrawal [23, 35], confirming that negative emotional responses are easily aroused under these circumstances.…”

“…USVs have received increased attention in drug abuse studies because administration of cocaine [18], amphetamine [19] and drug-associated cues [20, 21] increase 50–55 kHz frequency-modulated (FM) USV emissions. In addition, escalated levels of alcohol consumed by alcohol-dependent rats are significantly correlated with alcohol anticipatory 50–55 kHz FM USVs [22] and alcohol-dependent rats in a state of withdrawal are more easily provoked to emit negative affect-associated 22–28 kHz USVs by mild aversive stimuli [23, 24]. Ascending mesolimbic cholinergic [25] and dopaminergic pathways [26] mediate production of 22–28 kHz and 50–55 kHz FM USVs, repeatedly, in correspondence with negative and positive emotional states [14, 27, 28].…”

“…Additionally, we performed in-depth examinations of the acoustic properties of 22–28 kHz and 50–55 kHz FM USVs, including mean frequency and duration, with the prediction that these parameters will be altered by alcohol experience. We previously reported that alcohol-experienced P rats show decreased mean frequency of 22–28 kHz USVs compared to alcohol-naïve P rats, while call duration was unaltered in both USV types [24]. Other studies have shown that mean frequency and duration of 22–28 kHz USVs were altered under different drug treatments, but 50 kHz FM USV acoustic patterns remained constant [27, 38, 39].…”