Rationale
50-kHz ultrasonic vocalizations (USVs) emitted by adult rats are heterogeneous; they occur over a wide frequency range, show varying degrees of frequency modulation, and appear to differ in ...their behavioral significance. However, they have not been extensively categorized.
Objectives
The main objective of this study was to identify subtypes of 50-kHz USVs emitted by adult rats and to determine how amphetamine (AMPH) or social testing condition affects their relative and absolute production rate and acoustic characteristics. A second objective was to determine the extent of individual differences in call rate, call subtype profile, and acoustic parameters (i.e., duration, bandwidth, and mean peak frequency).
Methods
Adult male Long–Evans rats were administered systemic amphetamine (0.25–2 mg/kg, IP) and tested individually or with a cage mate for 20 min. Call categories were defined based on visual inspection of over 20,000 USV spectrograms. Surgical devocalization was performed on a subset of AMPH-tested rats in order to confirm the authenticity of call subtypes.
Results
Fourteen categories of 50-kHz USVs were recognized. Call subtypes were differentially affected by social context, AMPH dose, and time within session. In contrast, the acoustic characteristics of call subtypes were notably stable. Marked and stable inter-individual differences occurred with respect to overall 50-kHz call rate, acoustic parameters, and call profile.
Conclusions
The present findings, obtained under saline and amphetamine test conditions, provide the first detailed classification of adult rat 50-kHz USVs. Consideration of 50-kHz USV subtypes may advance our understanding of inter-rat communication and affective state.
Sucrose preference (SP) is a widely used measure of anhedonia in rat models of depression, yet depressed patients do not reliably show an analogous deficit. As an alternative affect-related measure, ...adult rat ultrasonic vocalizations (USVs) are attracting interest, but it is unclear whether SP and USVs provide independent measures. Here, we have assessed whether SP and USV emission are correlated in the absence of a depressogenic procedure. To this end, 24 male Long-Evans rats were tested daily for 24 days, with alternating SP tests and USV recordings; after a 3-month hiatus, USV emission was re-evaluated for 6 more days. SP was measured in simultaneous two-bottle choice tests, and USVs were recorded in an open field. The main measures were: SP, 50-kHz call rate, and relative prevalence of trill and flat call subtypes. These measures showed temporally-stable individual differences across the initial 24-day testing period, and at the 3-month USV follow-up tests. Correlational analysis revealed no significant relationships between SP and the three main USV measures. Rats differed consistently, not only in their 50-kHz call rates but also in their 50-kHz call profiles (i.e., the relative prevalence of 14 call subtypes); most rats preferentially emitted either trill or flat calls. Several inter-call subtype associations were detected, including a strong negative relationship between the relative prevalence of flat and trill calls. The 50-kHz call rate was correlated with the relative prevalence of only one call subtype (short calls, negative correlation), but was positively correlated with absolute emission rates for almost all subtypes. In conclusion, adult rats exhibited temporally-stable individual differences over weeks (SP) or months (USVs) of testing. This trait-like stability helped to reveal a lack of relationship between SP and the USV-related variables under study, suggesting that these measures may capture different constructs of possible relevance to animal models of depression.
The functional and molecular similarities and distinctions between human and murine astrocytes are poorly understood. Here, we report the development of an immunopanning method to acutely purify ...astrocytes from fetal, juvenile, and adult human brains and to maintain these cells in serum-free cultures. We found that human astrocytes have abilities similar to those of murine astrocytes in promoting neuronal survival, inducing functional synapse formation, and engulfing synaptosomes. In contrast to existing observations in mice, we found that mature human astrocytes respond robustly to glutamate. Next, we performed RNA sequencing of healthy human astrocytes along with astrocytes from epileptic and tumor foci and compared these to human neurons, oligodendrocytes, microglia, and endothelial cells (available at http://www.brainrnaseq.org). With these profiles, we identified novel human-specific astrocyte genes and discovered a transcriptome-wide transformation between astrocyte precursor cells and mature post-mitotic astrocytes. These data represent some of the first cell-type-specific molecular profiles of the healthy and diseased human brain.
•We develop the first method to acutely purify fetal and adult human astrocytes•We obtain transcriptome profiles of human neurons, glia, and vascular cells•While similar, human and mouse astrocytes have unique genomic and functional traits•Human astrocytes exist in at least two distinct developmental stages
Zhang et al. developed a method to acutely purify healthy and diseased human astrocytes and to culture them in serum-free conditions. They obtained transcriptome profiles of purified human CNS cell types and discovered two distinct stages of human astrocyte development.
Rationale
The behavioral effects of the nicotine metabolites nornicotine and cotinine have not been investigated extensively.
Objectives
To evaluate the effects of nicotine, cotinine, and ...nornicotine, given alone or in combination, on locomotor activity and emission of ultrasonic vocalizations in male adult rats.
Methods
Rats were first given home cage nicotine injections to make them tolerant to the drug’s locomotor depressant effects. On subsequent days, locomotor activity (LMA) and ultrasonic vocalizations were recorded in an open field, for 60 min after challenge injection, using repeated measures designs. In single-drug experiments, subjects were tested with nicotine 0.05–0.4 mg/kg, cotinine 0.03–3 mg/kg, or nornicotine 0.1–10 mg/kg. In drug-combination experiments, saline or nicotine 0.2 mg/kg challenge was preceded by cotinine (0, 0.3, 3 mg/kg) or nornicotine (0, 0.1, 0.3, 1, 3 mg/kg) injection.
Results
High doses of nornicotine increased LMA and blunted the locomotor stimulant effect of nicotine. Less consistently, nicotine and high doses of nornicotine decreased the 50-kHz call rate, with no clear evidence of a nornicotine × nicotine interaction. Cotinine, given alone or before nicotine injection, altered neither LMA nor the call rate. No drug altered the relative prevalence of flat vs. trill 50-kHz call subtypes, except that the highest dose of nornicotine promoted flat calls over trills. No drug evoked 22-kHz calls.
Conclusion
Nornicotine can exert an acute anti-nicotine effect in vivo
,
as previously reported in vitro. The finding that nicotine did not detectably alter the 50-kHz call profile appears consistent with this drug’s mild subjective effects in human subjects.
Background: Mechanisms underlying psychostimulant euphoria remain poorly understood. In adult rats, positive emotional states are associated with alterations in 50-kHz ultrasonic vocalizations ...(USVs): specifically, “trill” calls are promoted over “flat” calls. Here, we investigated the effects of acute and repeated cocaine administration, and—based on previous findings with amphetamine—their possible dependence on beta-adrenergic receptors. Methods: Adult male Long-Evans rats received intraperitoneal drug or saline injections before daily USV recording. Fourteen 50-kHz call subtypes were analyzed. In Experiments 1 and 2, cocaine (1–10 mg/kg) and propranolol (10 mg/kg) were tested alone. In Experiment 3, propranolol/cocaine interactions were sought within a conditioned place preference (CPP) procedure. Experiment 4 investigated acute and chronic cocaine effects (Phase 1), and propranolol/cocaine interactions either in an open field (Phase 2) or within a CPP procedure (Phase 3). Results: In drug-naïve animals, cocaine increased the 50-kHz call rate, with sensitization developing rapidly. After more extended exposure, cocaine now also increased the relative prevalence of trill versus flat calls; effects on other subtypes were also revealed. The beta-blocker propranolol prevented neither cocaine CPP nor cocaine effects on USV emission or locomotion but exerted significant USV-related effects when given alone. CPP magnitude and USV-related measures were uncorrelated. Conclusions: With long-term intraperitoneal administration, cocaine can alter the relative prevalence of several 50-kHz call subtypes; its ability to promote trill versus flat calls, in particular, is consistent with a positive affect interpretation. Cocaine’s behavioral effects (i.e., USV-related, locomotor, CPP) appear independent of beta-adrenergic receptor activity.
Rationale
Systemic amphetamine (AMPH) administration increases the rate of 50-kHz ultrasonic vocalizations (USVs) in adult rats and preferentially enhances the ‘trill’ subtype; these effects of AMPH ...critically depend on noradrenergic transmission, but the possible contributions of dopamine are unclear.
Objective
To assess the role of dopamine in 50-kHz USVs emitted drug-free and following systemic AMPH administration.
Methods
Adult male Long–Evans rats pre-selected for high AMPH-induced calling rates were tested with AMPH (1 mg/kg, intraperitoneal (IP)) and saline following pretreatment with the following dopamine receptor antagonists: SCH 23390 (0.005–0.02 mg/kg, subcutaneous (SC)), SCH 39166 (0.03–0.3 mg/kg, SC), haloperidol (0.1, 0.2 mg/kg, IP), sulpiride (20–80 mg/kg, SC), raclopride (0.1–0.5 mg/kg, SC), clozapine (4 mg/kg, SC), risperidone (0.5 mg/kg, SC), and pimozide (1 mg/kg, IP). The dopamine and noradrenaline reuptake inhibitors (GBR 12909 and nisoxetine, respectively) were also tested, alone and in combination.
Results
SCH 23390, SCH 39166, haloperidol, and raclopride dose-dependently inhibited vocalizations under AMPH and suppressed the proportion of trill calls. Sulpiride, however, had no discernable effect on call rate or profile, even at a high dose that reduced locomotor activity. Single doses of clozapine, risperidone, and pimozide all markedly decreased calling under saline and AMPH. Finally, GBR 12909 and nisoxetine failed to promote 50-kHz USVs detectably or alter the subtype profile, when tested alone or in combination.
Conclusions
The rate of 50-kHz USVs and the call subtype profile following systemic AMPH administration depends on dopaminergic neurotransmission through D1-like and D2-like receptors. However, inhibiting dopamine and/or noradrenaline reuptake appears insufficient to induce calling.
Rationale
Adult rat 22-kHz vocalizations are often associated with alarm or distress, whereas a subset of 50-kHz calls is preferentially emitted in response to amphetamine and other rewarding ...stimuli. Whether any 50-kHz calls reflect anxiety is unknown.
Objective
To determine the effects of anxiogenic drugs on 50-kHz call rate and call subtype profile, in comparison with
d
-amphetamine.
Methods
Adult male rats received systemic amphetamine (1 mg/kg) three times several days before testing. Ultrasonic vocalizations were then recorded after acute intraperitoneal injection of amphetamine or one of five anxiogenic drugs: yohimbine (2.5 mg/kg),
N
-methyl-
β
-carboline-3-carboxamide (FG 7142, 5 mg/kg), pentylenetetrazol (PTZ, 20 mg/kg),
m
-chlorophenylpiperazine (mCPP, 1 mg/kg), caffeine (25 mg/kg), or vehicle.
Results
The duration of immobility was increased by FG 7142, PTZ, and mCPP; this measure was unchanged by yohimbine and reduced by the locomotor stimulant drugs amphetamine and caffeine. Conversely, the 50-kHz call rate was reduced by FG 7142, PTZ and mCPP, and increased by caffeine and amphetamine. Overall, the most common 50-kHz call subtypes were flat, trill, step-up, and complex. Consistent with previous reports, amphetamine increased the relative prevalence of trill calls while reducing the relative prevalence of flat calls. Yohimbine and caffeine reduced flat call prevalence, whereas mCPP reduced trill call prevalence. No other shifts in the call profile were observed, and no anxiogenic drug induced 22-kHz calls.
Conclusion
Anxiogenic drugs, as a class, did not uniformly alter the 50-kHz call rate or subtype profile. Amphetamine-induced effects on 50-kHz call rate and profile do not reflect anxiety.
Rationale
Reinforcement-enhancing effects of nicotine occur in human subjects and laboratory rats. However, the doses used in animal studies typically exceed smoking-associated levels of exposure, ...and generalized behavioral activation by nicotine can potentially confound data interpretation.
Methods
During daily 60-min sessions, male adult rats pressed an “active” lever to illuminate a brief cue light. Pressing on either the active or inactive lever retracted both levers for 60 s. Nicotine (0.025–0.2 mg/kg) was given either by continuous intravenous (IV) infusion, or spaced IV pulses (3-s or 30-s/pulse), or pre-session subcutaneous (SC) injection.
Results
Almost all rats responded preferentially for the cue light for several weeks. After several home-cage nicotine injections, reinforcement enhancement occurred even within the first nicotine test session. Nicotine increased active lever responding without altering inactive lever responding, with effects reliably observed at doses as low as 0.1 mg/kg SC or 0.1 mg/kg/session IV. Within the session, the 0.1 mg/kg dose maximally increased active lever responding by 2–3-fold, coinciding with serum levels of 25 ng/ml. Intravenous nicotine (tested at 0.1 mg/kg/60-min session) was equally effective whether delivered by continuous infusion or in a series of equally spaced 0.003 mg/kg pulses each of 3-s or 30-s duration.
Conclusions
Low doses of nicotine can potentiate responding for a primary sensory reinforcer without producing a generalized increase in lever pressing. Reinforcer enhancement by nicotine generalized to several modes of drug delivery, appeared to track circulating levels of drug, and occurred even at serum levels within the daytime range of moderate cigarette smokers.
Rationale
Adult rats emit ultrasonic calls at around 22 and 50 kHz, which are often elicited by aversive and rewarding stimuli, respectively. Dopamine (DA) plays a role in aspects of both reward and ...aversion.
Objective
The purpose of this study is to investigate the effects of DA receptor subtype-selective agonists on 22- and 50-kHz call rates.
Methods
Ultrasonic calls were recorded in adult male rats that were initially screened with amphetamine to eliminate low 50-kHz callers. The remaining subjects were tested after acute intraperitoneal or subcutaneous injection of the following DA receptor-selective agonists and antagonists: A68930 (D1-like agonist), quinpirole (D2-like agonist), PD 128907 (D3 agonist), PD 168077 (D4 agonist), SCH 39166 (D1-like antagonist), L-741,626 (D2 antagonist), NGB 2904 (D3 antagonist), and L-745,870 (D4 antagonist). The indirect DA/noradrenaline agonist amphetamine served as a positive control.
Results
As expected, amphetamine strongly increased 50-kHz call rates. In contrast, D1-, D2-, and D3-selective DA receptor agonists, when given alone, inhibited calling; combinations of D1- and D2-like agonists also decreased call rate. Given alone, the D1-like and D3 antagonists significantly decreased call rate, with a similar trend for the D2 antagonist. Agonist–antagonist combinations also decreased calling. The D4 agonist and antagonist did not significantly affect 50-kHz call rates. Twenty-two-kilohertz calls occurred infrequently under all drug conditions.
Conclusion
Following systemic drug administration, tonic pharmacological activation of D1-like or D2-like DA receptors, either alone or in combination, does not appear sufficient to induce 50-kHz calls. Dopaminergic transmission through D1, D2, and D3 receptors appears necessary for spontaneous calling.
Rationale and objectives
The reinforcement-enhancing effect (REE) of nicotine refers to the drug’s ability to enhance the strength of other primary and conditioned reinforcers. The main aim was to ...investigate neuropharmacological mechanisms underlying nicotine’s strengthening of a primary visual reinforcer (i.e., a light cue), using a subcutaneous (SC) dose previously shown to provide plasma nicotine levels associated with habitual smoking.
Methods
Adult male rats pressed an “active” lever to illuminate a brief cue light during daily 60-min sessions. Rats that showed a clear REE were tested with systemically administered pretreatment drugs followed by nicotine (0.1 mg/kg SC) or saline challenge, in within-subject counterbalanced designs. Pretreatments were mecamylamine (nicotinic, 0.1-1 mg/kg SC), SCH 39166 (D1-like dopaminergic, 0.003-0.2 mg/kg SC), naloxone (opioid, 1 and 5 mg/kg SC), prazosin (alpha1-adrenergic antagonist, 1 and 2 mg/kg IP), rimonabant (CB1 cannabinoid inverse agonist, 3 mg/kg IP), sulpiride (D2-like dopaminergic antagonist, 40 mg/kg SC), or propranolol (beta-adrenergic antagonist, 10 mg/kg IP).
Results
The nicotine REE was abolished by three antagonists at doses that did not impact motor output, i.e., mecamylamine (1 mg/kg), SCH 39166 (0.01 and 0.03 mg/kg), and naloxone (5 mg/kg). Prazosin and rimonabant both attenuated the nicotine REE, but rimonabant also suppressed responding more generally. The nicotine REE was not significantly altered by sulpiride or propranolol.
Conclusions
In adult male rats, the reinforcement-enhancing effect of low-dose nicotine depends on nicotinic receptor stimulation and on neurotransmission via D1/D5 dopaminergic, opioid, alpha1-adrenergic, and CB1 cannabinoid receptors.
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