Diptendu Chatterjee

Acute and chronic alcohol dose: population differences in behavior and neurochemistry of zebrafish.

The zebrafish has been in the forefront of developmental genetics for decades and has also been gaining attention in neurobehavioral genetics. It has been proposed to model alcohol‐induced changes in human brain function and behavior. Here, adult zebrafish populations, AB and SF (short‐fin wild type), were exposed to chronic treatment (several days in 0.00% or 0.50% alcohol v/v) and a subsequent acute treatment (1 h in 0.00%, 0.25%, 0.50% or 1.00% alcohol). Behavioral responses of zebrafish to computer‐animated images, including a zebrafish shoal and a predator, were quantified using videotracking. Neurochemical changes in the dopaminergic and serotoninergic systems in the brain of the fish were measured using high‐precision liquid chromatography with electrochemical detection. The results showed genetic differences in numerous aspects of alcohol‐induced changes, including, for the first time, the behavioral effects of withdrawal from alcohol and neurochemical responses to alcohol. For example, withdrawal from alcohol abolished shoaling and increased dopamine and 3,4‐dihydroxyphenylacetic acid in AB but not in SF fish. The findings show that, first, acute and chronic alcohol induced changes are quantifiable with automated behavioral paradigms; second, robust neurochemical changes are also detectable; and third, genetic factors influence both alcohol‐induced behavioral and neurotransmitter level changes. Although the causal relationship underlying the alcohol‐induced changes in behavior and neurochemistry is speculative at this point, the results suggest that zebrafish will be a useful tool for the analysis of the biological mechanisms of alcohol‐induced functional changes in the adult brain.

High precision liquid chromatography analysis of dopaminergic and serotoninergic responses to acute alcohol exposure in zebrafish

Zebrafish is gaining popularity in behavioral neuroscience in general and in alcohol research in particular. Alcohol is known to affect numerous molecular mechanisms depending on dose and administration regimen. Prominent among these mechanisms are several neurotransmitter systems. Here we analyze the responses of the dopaminergic and serotoninergic neurotransmitter systems of zebrafish to acute alcohol treatment (1 h long exposure of adult fish to 0.00%, 0.25%, 0.50%, or 1.00% ethyl alcohol) by testing the concentration of dopamine, its metabolite DOPAC, and serotonin and its metabolite 5-HIAA from whole brain extracts. We utilize a sensitive HPLC method and describe significant alcohol induced changes in zebrafish for the first time. We show that dopamine significantly increased in a quasi-linear dose dependent manner, DOPAC showed a smaller apparent increase which was non-significant, while both serotonin and 5-HIAA showed a significant increase only in the highest acute dose group. We discuss the methodological novelty of our work and theorize about the implications of the neurotransmitter level changes from a behavioral perspective.

Hormones that increase maternal responsiveness affect accumbal dopaminergic responses to pup- and food-stimuli in the female rat

The present study investigated hormonal mediation of maternal behavior and accumbal dopamine (DA) responses to pup-stimuli, as measured in microdialysis samples collected from the nucleus accumbens shell of female rats in non-homecage environment. In Experiment 1, samples were collected before and after continuous homecage pup experience from either intact postpartum or cycling females. In Experiment 2, samples were collected before and after responding maternally in homecage from ovariectomized females given either parturient-like hormone or sham treatments. After baseline sample collection in the dialysis chamber, pup and food stimuli were individually presented to females. Upon sampling completion, all animals were placed back into their homecage with donor pups for several days, and then the sample collection procedure was repeated. Prior to stimulus presentation, postpartum and hormone-treated females had decreased basal DA release compared to their controls. In response to pup stimuli, only postpartum and hormone-treated females had increased DA release compared to basal release (both sampling days). In response to food stimuli, all females had increased DA responses from basal; although there were group differences on the initial day of sampling. Findings suggest that hormones associated with inducing maternal behavior in the postpartum rat play a significant role in modifying accumbal dopaminergic responses on first exposure to pup stimuli in the rat. However, the postpartum experience provides further modifications to this brain region to promote DA responses to pup stimuli.

Previous maternal experience affects accumbal dopaminergic responses to pup-stimuli

The present study investigated the release of dopamine from the nucleus accumbens (shell) in response to pup-stimuli in the absence of lactation and maternal behaviors at time of sample collection. Subjects were female rats given maternal experiences through prior parturitions, recent pup-induced sensitization, or a combination of both. Nulliparous (N) or multiparous (M, had 2 prior litters but cycling) female rats either received pup-sensitization (S+) until they responded maternally in their homecage or no pup-sensitization (S-), thus, there were four groups: NS- (n=5), NS+ (n=6), MS- (n=5), and MS+ (n=8). Four hours after removal of pups (from homecage for S+ groups), all females were placed into the microdialysis chamber for sample collection. After baseline collection, four foster pups were given to the females. In this paradigm females show little to no maternal behavior in the test chamber. Samples (collected every 8 min) were analyzed for dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) with electrochemical detection using HPLC. Relative to the inexperienced NS- females, the experienced NS+, MS- and MS+ females displayed significantly increased DA levels only during the first 8 min of pup-exposure. The more experience a female had with pups, the greater was the DA response (p<.05). The results suggest that enhanced responding to pups following previous maternal experiences may be mediated through accumbal DA.

Dopamine receptor antagonism disrupts social preference in zebrafish: a strain comparison study.

Zebrafish form shoals in nature and in the laboratory. The sight of conspecifics has been found reinforcing in zebrafish learning tasks. However, the mechanisms of shoaling, and those of its reinforcing properties, are not known. The dopaminergic system has been implicated in reward among other functions and it is also engaged by drugs of abuse as shown in a variety of vertebrates including zebrafish. The ontogenetic changes in dopamine levels and, to a lesser degree, in serotonin levels, have been found to accompany the maturation of shoaling in zebrafish. Thus, we hypothesized that the dopaminergic system may contribute to shoaling in zebrafish. To test this we employed a D1-receptor antagonist and quantified behavioral responses of our subjects using a social preference (shoaling) paradigm. We found significant reduction of social preference induced by the D1-R antagonist, SCH23390, in the AB strain of zebrafish, an alteration that was not accompanied by changes in motor function or vision. We also detected D1-R antagonist-induced changes in the level of dopamine, DOPAC, serotonin and 5HIAA, respectively, in the brain of AB zebrafish as quantified by HPLC with electrochemical detection. We found the antagonist-induced behavioral changes to be absent and the levels of these neurochemicals to be lower in another zebrafish population, SF, demonstrating naturally occurring genetic variability in these traits. We conclude that this variability may be utilized to unravel the mechanisms of social behavior in zebrafish, a line of research that may be extended to other vertebrates including our own species.

Maturation of shoaling in two zebrafish strains: a behavioral and neurochemical analysis.

Abnormal social behavior is a hallmark of several human neuropsychiatric and neurodevelopmental disorders for which appropriate treatment is lacking. The zebrafish has been proposed as a tool with which these disorders may be modeled and their mechanisms analyzed. A potential starting point of such analyses is the identification of genetic differences between distinct zebrafish strains. Here we compare AB and TU, two well established zebrafish strains, and characterize the developmental trajectories of their shoaling (social) behavior and of the levels of dopamine, serotonin as well as a metabolite of each of these neurotransmitters, DOPAC and 5HIAA from whole brain extracts. Using a novel video-tracking software application, we demonstrate significant strain dependent changes in the maturation of shoaling between day 7 and day 87 post-fertilization. Using high-precision liquid chromatography specifically adapted to zebrafish, we uncover a significant age×strain interaction in dopamine and DOPAC that apparently correlates well with the behavioral differences found between the strains. We also report on strain differences in serotonin and 5HIAA. We discuss possible mechanistic analyses that will address causality and conclude that zebrafish will be a useful tool with which the neurobiological and genetic bases of social behavior may be analyzed in vertebrates.

Sight of conspecific images induces changes in neurochemistry in zebrafish.

Zebrafish are gaining popularity in behavioural brain research as this species combines practical simplicity with system complexity. The dopaminergic system has been thoroughly investigated using mammals. Dopamine plays important roles in motor function and reward. Zebrafish have dopamine receptors homologous to mammalian counterparts, and dopamine receptor antagonists as well as alcohol have been shown to exert significant effects on this species as measured using HPLC or behavioural methods. The sight of conspecifics was previously shown to be rewarding in zebrafish but whether this stimulus affects the dopaminergic system has not been studied. Here, we present animated images of zebrafish to the experimental zebrafish subject for varying lengths of time and quantify the amount of dopamine, DOPAC, serotonin and 5HIAA extracted from the subjects brain immediately after the stimulus presentation using HPLC with electrochemical detection. We find conspecific images to induce a robust behavioural response (attraction) in experimental zebrafish. Importantly, dopamine and DOPAC levels significantly increased in response to the presentation of conspecific images but not to scrambled images. Last, serotonin and 5HIAA levels did not significantly change in response to the conspecific images. We conclude that our findings, together with pervious studies, now conclusively demonstrate that the behavioural response induced by the appearance of conspecifics is mediated, at least partly, by the dopaminergic system in zebrafish.

Experience-dependent cell survival in the maternal rat brain.

Postpartum maternal experience produces long-lasting changes in maternal behavior in the mother rat, which can be altered by early-life isolation. Postpartum experience also affects the regulation of adult neurogenesis in the neural circuit underlying maternal behavior, in a region-specific manner. Female rats were reared either with their mothers (MR) or in isolation in an artificial rearing (AR) paradigm. In adulthood, rats were mated and separated from their pups at birth. The following day, dams were injected with a mitotic marker and either allowed to interact with pups (maternal experience) or left alone. Results show that MR rats that acquire a later maternal experience show increases in cell survival in parts of the excitatory limb of the maternal neural network (bed nucleus of the stria terminalis and nucleus accumbens), but no changes in the inhibitory limb (amygdala). In comparison to AR inexperienced rats, AR maternally experienced rats show no increases in cell survival in the excitatory limb, but a striking reduction in cell survival in the inhibitory limb. The results suggest that early preweaning maternal isolation alters the structural plasticity that occurs following a postpartum maternal experience.

An integrative analysis of ethanol tolerance and withdrawal in zebrafish (Danio rerio)

The zebrafish is emerging as a popular animal model for alcohol (ethanol or EtOH) addiction due to its simplicity and practical advantages. Two phenomena associated with ethanol addiction are the development of tolerance and withdrawal. Using a multi-level approach in the current study, we characterise ethanol tolerance and withdrawal in zebrafish. We first investigate the temporal trajectory of ethanol concentration in the zebrafish brain in response to an acute exposure and during withdrawal. We report that ethanol concentrations approach a steady state within 60 min of exposure to 0.50% and 1.00% v/v ethanol and rapidly decline and return to zero within 60 min following withdrawal from chronic ethanol exposure (0.50% v/v). We characterise the changes associated with ethanol tolerance and withdrawal in zebrafish by focusing on three domains relevant to ethanol addiction: motor patterns, physiological responses (i.e. cortisol levels), and neurochemical alterations. The use of multiple domains of investigation allowed an in-depth analysis of ethanol induced changes in zebrafish.

Myocyte Androgen Receptors Increase Metabolic Rate and Improve Body Composition by Reducing Fat Mass

Testosterone and other androgens are thought to increase lean body mass and reduce fat body mass in men by activating the androgen receptor. However, the clinical potential of androgens for improving body composition is hampered by our limited understanding of the tissues and cells that promote such changes. Here we show that selective overexpression of androgen receptor in muscle cells (myocytes) of transgenic male rats both increases lean mass percentage and reduces fat mass. Similar changes in body composition are observed in human skeletal actin promoter driving expression of androgen receptor (HSA-AR) transgenic mice and result from acute testosterone treatment of transgenic female HSA-AR rats. These shifts in body composition in HSA-AR transgenic male rats are associated with hypertrophy of type IIb myofibers and decreased size of adipocytes. Metabolic analyses of transgenic males show higher activity of mitochondrial enzymes in skeletal muscle and increased O(2) consumption by the rats. These results indicate that androgen signaling in myocytes not only increases muscle mass but also reduces fat body mass, likely via increases in oxidative metabolism.