"C.R.E.A.T.E."-ing Unique Primary-Source Research Paper Assignments for a Pleasure and Pain Course Teaching Neuroscientific Principles in a Large General Education Undergraduate Course.

A large (250 registrants) General Education lecture course, Pleasure and Pain, presented basic neuroscience principles as they related to animal and human models of pleasure and pain by weaving basic findings related to food and drug addiction and analgesic states with human studies examining empathy, social neuroscience and neuroeconomics. In its first four years, the course grade was based on weighted scores from two multiple-choice exams and a five-page review of three unique peer-reviewed research articles. Although well-registered and well-received, 18% of the students received Incomplete grades, primarily due to failing to submit the paper that went largely unresolved and eventually resulted in a failing grade. To rectify this issue, a modified version of the C.R.E.A.T.E. (Consider, Read, Elucidate hypotheses, Analyze and interpret data, Think of the next Experiment) method replaced the paper with eight structured assignments focusing on an initial general-topic article, the introduction-methods, and results-discussion of each of three related peer-review neuroscience-related articles, and a final summary. Compliance in completing these assignments was very high, resulting in only 11 INC grades out of 228 students. Thus, use of the C.R.E.A.T.E. method reduced the percentage of problematic INC grades from 18% to 4.8%, a 73% decline, without changing the overall grade distribution. Other analyses suggested the students achieved a deeper understanding of the scientific process using the C.R.E.A.T.E. method relative to the original term paper assignment.

Pleasure and pain: teaching neuroscientific principles of hedonism in a large general education undergraduate course.

In a large (250 registrants) general education lecture course, neuroscience principles were taught by two professors as co-instructors, starting with simple brain anatomy, chemistry, and function, proceeding to basic brain circuits of pleasure and pain, and progressing with fellow expert professors covering relevant philosophical, artistic, marketing, and anthropological issues. With this as a base, the course wove between fields of high relevance to psychology and neuroscience, such as food addiction and preferences, drug seeking and craving, analgesic pain-inhibitory systems activated by opiates and stress, neuroeconomics, unconscious decision-making, empathy, and modern neuroscientific techniques (functional magnetic resonance imaging and event-related potentials) presented by the co-instructors and other Psychology professors. With no formal assigned textbook, all lectures were PowerPoint-based, containing links to supplemental public-domain material. PowerPoints were available on Blackboard several days before the lecture. All lectures were also video-recorded and posted that evening. The course had a Facebook page for after-class conversation and one of the co-instructors communicated directly with students on Twitter in real time during lecture to provide momentary clarification and comment. In addition to graduate student Teaching Assistants (TAs), to allow for small group discussion, ten undergraduate students who performed well in a previous class were selected to serve as discussion leaders. The Discussion Leaders met four times at strategic points over the semester with groups of 20-25 current students, and received one credit of Independent Study, thus creating a course within a course. The course grade was based on weighted scores from two multiple-choice exams and a five-page writing assignment in which each student reviewed three unique, but brief original peer-review research articles (one page each) combined with expository writing on the first and last pages. A draft of the first page, collected early in the term, was returned to each student by graduate TAs to provide individual feedback on scientific writing. Overall the course has run three times at ful or near enrollment capacity despite being held at an 8:00 AM time slot. Student-generated teaching evaluations place it well within the normal range, while this format importantly contributes to budget efficiency permitting the teaching of more required small-format courses (e.g., freshman writing). The demographics of the course have changed to one in which the vast majority of the students are now outside the disciplines of neuroscience or psychology and are taking the course to fulfill a General Education requirement. This pattern allows the wide dissemination of basic neuroscientific knowledge to a general college audience.

Differential activation of cAMP response element binding protein in discrete nucleus accumbens subregions during early and late cocaine sensitization.

The present study examined the differential cocaine-induced activation of the cyclic adenosine monophosphate (cAMP) response element binding protein (CREB) throughout discrete zones of analysis of the nucleus accumbens (NAc) in rats. CREB-dependent gene transcription, which may underlie long-lasting drug-induced changes in behavior and the subjective effects of cocaine, varies depending on the stage of drug exposure or withdrawal and the cell population involved. Using immunohistochemistry, the authors analyzed changes in CREB phosphorylation in the NAc after 5 days of cocaine, a short or long drug-free period, and a subsequent challenge injection. The NAc shell was separated into 5 zones of analysis previously defined by neurochemistry and connectivity. Repeated cocaine resulted in CREB phosphorylation in all analyzed subregions of the NAc excluding the most ventrolateral region of the shell 2 weeks after cessation of repeated cocaine, but rats challenged after 2 drug-free days yielded a more localized activation of CREB in the 3 most dorsomedial zones of the shell. The temporal and anatomical determinants of cocaine-induced CREB activity may indicate functional differences among NAc shell subregions and suggest the involvement of CREB in early and late cocaine effects.

Electrolytic lesions of a discrete area within the nucleus accumbens shell attenuate the long-term expression, but not early phase, of sensitization to cocaine.

Repeated exposure to cocaine leads to behavioral sensitization, which is the augmentation of the locomotor response to a subsequent exposure to the drug. The nucleus accumbens (NAc), a major termination site of dopaminergic neurons, is believed to be involved in behavioral sensitization and studies have demonstrated that the NAc shell can be split into five zones of analysis; the vertex, arch, cone, intermediate and ventrolateral zones [Todtenkopf MS, Stellar JR. Assessment of tyrosine hydroxylase immunoreactive innervation in five subregions of the nucleus accumbens shell in rats treated with repeated cocaine. Synapse 2000;38:261-70]. Several reports show cocaine-induced c-fos expression particularly in the intermediate zone after 14, but not 2, drug-free days following repeated cocaine administration, suggesting that this region may be involved in sensitization and particularly in the later phase of expression, versus the earlier phase of sensitization. Bilateral electrolytic lesions of the intermediate zone were made in two groups of rats, which were then repeatedly exposed to cocaine (15 mg/kg, twice/day for 5 days). One group was subsequently given a single cocaine challenge injection (15 mg/kg) after 14 drug-free days, while the other group was challenged after only 2 drug-free days. Two sham surgery groups in which an electrode was lowered but no current was passed served as controls. Results show that lesioned animals as well as sham controls exhibited behavioral sensitization to the drug. However, following a 14-day drug-free period, the lesioned animals showed significant reduction in sensitization, compared to sham controls. Together these findings suggest that the intermediate zone of the NAc shell is indeed involved in the expression phase of behavioral sensitization to cocaine.

Assessing contributions of nucleus accumbens shell subregions to reward-seeking behavior.

BACKGROUND: The nucleus accumbens (NAc) plays a key role in brain reward processes including drug seeking and reinstatement. Several anatomical, behavioral, and neurochemical studies discriminate between the limbic-associated shell and the motor-associated core regions. Less studied is the fact that the shell can be further subdivided into a dorsomedial shell (NAcDMS) and an intermediate zone (NAcINT) based on differential expression of transient c-Fos and long-acting immediate-early gene ΔFosB upon cocaine sensitization. These disparate expression patterns suggest that NAc shell subregions may play distinct roles in reward-seeking behavior. In this study, we examined potential differences in the contributions of the NAcDMS and the NAcINT to reinstatement of reward-seeking behavior after extinction. METHODS: Rats were trained to intravenously self-administer cocaine, extinguished, and subjected to a reinstatement test session consisting of an intracranial microinfusion of either amphetamine or vehicle targeted to the NAcDMS or the NAcINT. RESULTS: Small amphetamine microinfusions targeted to the NAcDMS resulted in statistically significant reinstatement of lever pressing, whereas no significant difference was observed for microinfusions targeted to the NAcINT. No significant difference was found for vehicle microinfusions in either case. CONCLUSION: These results suggest heterogeneity in the behavioral relevance of NAc shell subregions, a possibility that can be tested in specific neuronal populations in the future with recently developed techniques including optogenetics.

Social stress does not alter the expression of sensitization to cocaine.

The effects of chronic social stress on behavioral sensitization to cocaine were investigated in the Syrian hamster. Adolescent animals received either 15 mg/kg i.p. of cocaine or saline twice per day for 7 consecutive days. Two weeks following the last injection (young adulthood), they were given a challenge dose of 5 mg/kg i.p. of cocaine and scored for locomotion. Motor activity was significantly greater in cocaine-treated animals, demonstrating sensitization to this psychostimulant. Following the results of the first study, another group of adolescent animals was exposed to either a novel clean cage (control) or an aggressive resident male hamster (social stress) for 15 min following an injection of cocaine (20 mg/kg i.p. once daily) or saline for 7 consecutive days. The groups were as follows: Social Stress/Cocaine (SSC), No Social Stress/Cocaine (NSSC), Social Stress/Saline (SSS) and No Social Stress/Saline (NSSS). Two weeks following the last injection (Day 21), all animals were given a challenge dose of cocaine (5 mg/kg i.p.) and were rescored for locomotion. At that time, the suppressive effect of stress on locomotion was no longer detectable, as the expression of sensitization was observed in the NSSC but not in the SSC group. These results suggest that chronic social stress administered during adolescence does not cross-sensitize with cocaine in young adult hamsters.

Both the shell of the nucleus accumbens and the central nucleus of the amygdala support amphetamine self-administration in rats.

Intracranial self-administration of drugs offers the opportunity to localize the neuronal substrates mediating the rewarding effects of drugs. The purpose of the present study was to explore whether the nucleus accumbens shell and the central nucleus of the amygdala, two components of the "extended amygdala," would support self-administration of the psychostimulant amphetamine. Male Wistar rats were trained to lever press under a Fixed Ratio 1 schedule of reinforcement for D-amphetamine injections (0, 1.0, 1.5, 2.0 and 3.0 microg/microl/inj) into either the nucleus accumbens shell or the central nucleus of the amygdala. An ascending limb dose-response function with peak responding at the 2.0 microg/microl/inj dose was obtained for self-administration at both brain sites. These results indicate that monoaminergic transmission in both the nucleus accumbens shell and the central nucleus of the amygdala mediates the rewarding effects of amphetamine. Further, the present study provides additional evidence about the functional homogeneity of the forebrain continuum called the "extended amygdala."

Subregion-specific down-regulation of 5-HT3 immunoreactivity in the nucleus accumbens shell during the induction of cocaine sensitization.

Repeated exposure to psychostimulants such as cocaine and amphetamine can result in behavioral sensitization, which is believed to model the onset of drug addiction, as well as neural adaptations that occur after repeated drug abuse that lead to addictive behaviors. Dopamine (DA) in the nucleus accumbens (NAc) has been shown to play an integral role in this phenomenon. However, cocaine also acts on the serotonin (5-HT) system, which has been shown to modulate psychostimulant-induced increases in motor behavior and DA release in the NAc. Recently, it has been demonstrated that the shell portion of the NAc can no longer be considered a homogeneous structure and can be subdivided into at least five separate regions. The present study examines 5-HT(3) receptors in the subdivisions of the NAc in cocaine-sensitized rats. Rats received a sensitization-inducing regimen of cocaine (twice-daily injections of 15 mg/kg ip for five consecutive days). Two or 14 days following the last injection, rats were given a challenge injection of cocaine (15 mg/kg ip) and sacrificed 2 h later. Sections of the NAc were processed for 5-HT(3) immunoreactivity (5-HT(3)-IR), and the number of puncta was quantified in each of the subregions of the shell, as well as the core of the accumbens. Repeated cocaine administration resulted in robust sensitization that correlated with a transient decrease in the density of 5-HT(3) immunoreactive puncta in the intermediate zone of the accumbens shell. After a 2-week withdrawal period, sensitized animals no longer showed any differences in any of the areas examined. These data suggest a possible role for 5-HT(3) receptors in the intermediate zone during the induction of cocaine sensitization.

Neural processing of a cocaine-associated odor cue revealed by functional MRI in awake rats.

Using an olfactory conditioning procedure, brain stimulation reward threshold measurements, and functional magnetic resonance imaging (fMRI), we investigated brain stimulation reward threshold change and fMRI neural activation in response to a cocaine-associated odor cue. In the first brain stimulation experiment, over 10 days of rate-frequency curve-shift testing, rats were administered intravenous cocaine (1.0mg/kg) paired with a contextual cue of peppermint odor previously placed in the operant chamber or they were given vehicle treatment (no cocaine) in the presence of no olfactory cue. Following a 14-day drug-free rest period, rats were again given the rate-frequency curve-shift threshold test with or without the odor cue. In a second experiment, rats were similarly conditioned with a peppermint odor but with intraperitoneally delivered cocaine (10mg/kg). After a 14 day rest period, rats were imaged on a 7-T MRI for their blood oxygen level dependent (BOLD) in response to the cocaine-paired peppermint odor versus an unpaired neutral lemon odor. In the brain stimulation experiment, expected significant reward threshold shifts were produced by cocaine and, importantly, about half that level of shift was produced by the paired contextual olfactory cue. In the fMRI experiment, the insular cortex showed a significantly greater BOLD activation in cocaine-treated versus saline-treated animals to the olfactory cue, but not with the unpaired lemon scent. These data are in agreement with previous studies suggesting a role of the insular cortex in attributing reward value (positive or negative) to conditioned odor stimuli.

A flexible system for hands-free intracranial microinjection.

Neuroscience research projects often use intracranial (IC) microinfusions to target drug delivery to specific brain areas during behavioral testing. These experiments require accurate and precisely-timed delivery of small volumes. We present here a stepper motor-powered micropump assembly for such delivery. This system is hands-free, does not use a potentially leaky fluid swivel or use long delivery tubes that are subject to peristaltic forces during animal movements, and has been applied in combination with other paradigms. This micropump system reliably delivers a wide range of fluid volumes (e.g. 50 nL to 1 microL in tissue or greater for intraventricular injections) bilaterally from two independent, commercially available microsyringes through standard surgically implanted guide cannulae. It is easy to build and disassemble for cleaning or changing microsyringes. This system can also be used for a variety of purposes, e.g. intracranial self-administration, place conditioning, and many more, with the advantage that it provides a way to gather important data in the seconds and minutes following IC microinfusion without disruption of the animal's behavior by handling.

Effects of individual and concurrent stimulation of striatal D1 and D2 dopamine receptors on electrophysiological and behavioral output from rat basal ganglia.

Bilateral infusions of d-amphetamine into the rat ventral-lateral striatum (VLS) were previously shown to cause a robust behavioral activation that was correlated temporally with a net increase in firing of substantia nigra pars reticulata (SNpr) neurons, a response opposite predictions of the basal ganglia model. The current studies assessed the individual and cooperative contributions of striatal D1 and D2 dopamine receptors to these responses. Bilateral infusions into VLS of the D1/D2 agonist apomorphine (10 microg/microl/side) caused intense oral movements and sniffing, and an overall increase in SNpr cell firing to 133% of basal rates, similar to effects of d-amphetamine. However, when striatal D2 receptors were stimulated selectively by infusions of quinpirole (30 microg/microl/side) + the D1 antagonist R-(+)-7-chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine (SCH 23390; 10 microg/microl/side), no behavioral response and only modest and variable changes in SNpr cell firing were observed. Selective stimulation of striatal D1 receptors by (+/-) 6-chloro-APB hydrobromide (SKF 82958; 10 microg/microl/side) + the D2 antagonist cis-N-(1-benzyl-2-methyl-pyrrolidin-3-yl)-5-chloro-2-methoxy-4-methyl-aminobenzamide (YM 09151-2; 2 microg/microl/side) caused a weak but sustained increase in oral movements and modestly increased SNpr cell firing, but neither response was of the magnitude observed with apomorphine. When the two agonists were infused concurrently, however, robust oral movements and sniffing again occurred over the same time period that a majority of SNpr cells exhibited marked, sometimes extreme and fluctuating, changes in firing (net increase, 117% of basal rates). These data confirm that concurrent striatal D1/D2 receptor stimulation elicits a strong motor activation that is correlated temporally with a net excitation rather than inhibition of SNpr firing, and reveal that D1 and D2 receptors interact synergistically within the striatum to stimulate both forms of output.