Investigating State-Switching to Egestive Motor Output Evoked by Arginine Vasotocin (AVT) in the Feeding System of the Pond Snail, Helisoma trivolvis

Chris Callahan (Neuroscience)

Advisor: Nancy Kleckner

The feeding central pattern generator (CPG) in the pond snail Helisoma trivolvis is composed of three groups of interneurons, S1, S2 & S3.  These interneurons innervate associated motor neurons that control protraction, retraction and hyper-retraction, respectively, of the tongue like radula.  The CPG is further modulated by Buccal A Cluster (BAC) neurons, a subset of heterogeneous electrotonically coupled cells located in the buccal ganglia.  In previous research, arginine vasotocin (AVT), neuropeptide phenylalanine (NPF) and gonatropin releasing hormong (GnRH) have been shown to evoke state-switching responses that implicate this BAC circuitry in the initiation of behavior such as regurgitation, satiation and substrate-cleaning.  The purpose of this study is to investigate the mechanisms controlling egestive behavior through simultaneous recordings in BAC neurons and S1/S3 motor neurons, as well as through immunohistochemical staining.  Findings suggest that AVT consistently evokes BAC firing, which is coupled to S2 and non-S2 IPSPs in phase 3 motor neuron B19.  Findings also suggest that AVT may act directly on the CPG neurons to evoke patterned motor output.  Staining for arginine vasopressin (AVP) revealed positive immunoreactivity in the neuropile of the buccal ganglia, the gastric nerve of the esophagus and the central ring of the CNS.  While findings are somewhat preliminary, these results bolster an argument for the presence of AVT in Helisoma trivolvis and its role in the initiation of egestive feeding rhythms.

Alexithymia and Working Memory: Performances on a Recency Probes WM Task   

Sean M. Colligan (Neuroscience)

Advisor: Nancy Koven

Alexithymia is a personality construct characterized by a deficit in emotional processing skills.  Recent evidence suggests that there may be cognitive deficits associated with alexithymia as well.  The current study was conducted to determine to the role that emotional stimuli may have on the cognitive abilities of alexithymics compared to non-alexithymics.  Participants (N = 93) completed the TAS-20 self-report questionnaire to assess their levels of alexithymia and a recency-probes working memory task to assess working memory skills.  In the recency-probes task, participants responded to words of positive, negative, and neutral valences, with some trials containing proactive interference from recent previous trials.  It has been shown that both emotionally valenced words may facilitate interference resolution compared to neutral words.  It was expected that alexithymic participants, with their impaired emotional processing, would show less interference resolution on the emotional trials.  Surprisingly, high alexithymics (n = 19) showed a complete lack of interference on trials with positive stimuli, while low alexithymics (n = 19) showed significantly more interference on these trials.  Additionally, positive correlations were found between TAS-20 scores and interference on trials with negative stimuli.  This contrasting relationship between emotional stimuli and interference in working memory suggests that alexithymics may be over-attending to negative stimuli while simultaneously under-attending to positive stimuli.  This may be explained by a decrease in left compared to right prefrontal cortex activity in these individuals.

Characterization of S1 Interneurons and Determination of their Roles in the Modulation of Phase 2 and 3 of the Pond Snail, Helisoma trivolvis Feeding Pattern

Deniz Nesli Dolcen  (Biology)

Advisor: Nancy Kleckner

This thesis focused on the modulation of the feeding CPG in the pond snail, Helisoma trivolvis. It has been shown that the S1 interneurons N1a and dopaminergic BCN1 exhibit prolonged action potential bursts during phase 1 in response to emetic stimuli, and this prolonged phase 1 activity has been shown to correlate with the strong protraction cycles seen during regurgitation. The role of these neurons in the modulation of phase 1 of the feeding pattern has been investigated; however, there has been no research on the role of these neurons in the modulation of phase 2 and 3 of the feeding pattern. In addition, the GABAergic BCN1s have been implicated in the elimination of the hyperretraction phase during the switch from ingestion to regurgitation; however, it has not been demonstrated that the stimulation of these neurons induce inhibitory inputs in S3 motor neurons during phase 3 of the feeding pattern to eliminate the hyperretraction of the odontophore. Therefore, this project aimed to determine the differential roles of N1a and the dopaminergic BCN1 in the modulation of the phase 2 and 3 of the Helisoma feeding pattern, test the validity of the proposed role for the GABAergic BCN1s and further characterize the interactions of all of these neurons in the Helisoma CNS. The electrical activities of putative N1a and BCN1 neurons were recorded and manipulated to observe the corresponding effects on the electrical activities of S2 or S3 motor neurons. The putative S1 interneurons were then stained with a fluorescent dye and processed with GAD65 antibodies to confirm their identities and determine whether they were GABAergic in order to analyze the correlation between the electrical activities of specific S1 interneurons and the motor neurons. Firing of the putative N1a alone under dopamine perfusion was enough to initiate and maintain the standard feeding pattern and was necessary for the stimulation of the S2 interneuron. NPF, which has been implicated in the switch to regurgitation, initiated the standard pattern in a putative N1a neuron and later switched the firing pattern into a faster S1-S2 rhythm, which is characteristic of regurgitation. Firing of the non-GABAergic BCN1 was not necessary for the activation of the S2 interneuron, suggesting that the dopaminergic BCN1 might be functioning in regulating the frequency of the feeding pattern. In addition, stimulation of the non-GABAergic BCN1 activated the feeding CPG and caused an S1-S2 rhythm in the S3 motor neuron B19, suggesting that the dopaminergic BCN1 might also be involved in the initiation of the regurgitation rhythm, perhaps by stimulating the GABAergic BCN1 neurons. Stimulation of a putatively GABAergic BCN1 neuron was necessary for the firing of the S2 interneuron, suggesting that the firing of the GABAergic BCN1 neurons might be necessary in the initiation and maintenance of the standard pattern through stimulating the dopaminergic S1 interneurons.

The Potential of NMDA Antagonist, MK-801, to Disrupt Reconsolidation and Reinstatement of a Morphine Withdrawal-Induced Conditioned Place Aversion

Kristen Finn (Neuroscience)

Advisor: John Kelsey

A commonly used treatment for drug addiction, known as extinction therapy, presents drug-associated cues in the absence of the drug in order to break the connection between the cues and the effect of the drug. However, subsequent exposure to stress, drug-cues, and the original context often reinstate the original drug-seeking behavior because extinction only suppresses the CS-US memory, rather than eliminating it. Consistent with this, in my first experiment, I showed that a morphine withdrawal-induced conditioned place aversion (CPA) could be extinguished, but that it could be reinstated by the stress of an injection of the opiate antagonist, naloxone. Recent studies have tried to more permanently eliminate a memory by using drugs to block reconsolidation. For example, it has been shown that the reactivation of a well-consolidated fear-associated memory causes it to enter a labile state during which it can be altered or completely eliminated with the use of amnesic drugs. Consistent with this, in my second experiment, I found that injections of the glutamatergic NMDA receptor antagonist MK-801 immediately after a series of re-exposures to the test apparatus designed to reactivate the CPA increased the loss of that aversion. However, the CPA was never completely eliminated. Moreover, I also found that MK-801 injections given 2 hr after CPA reactivation slowed the loss of the aversion, indicating that the role of NMDA receptors in reconsolidation and extinction is complex.

Effects of Electrolytic Lesions of the Medial Habenula on a Nicotine-Induced Conditioned Taste Aversion in Rats   

 Luke H. Harmeling (Psychology)

Advisor: John Kelsey

Nicotine has aversive effects which are noticeably perceived at high doses.  The medial habenula (MHb), which has nicotinic acetylcholine receptors, has been implicated in mediating these aversive effects.  To test this hypothesis, I determined if electrolytic lesions of the MHb would reduce a conditioned taste aversion (CTA) created by pairing intake of a novel Kool-Aid flavor with an i.p. injection of 0.8 mg/kg nicotine. It was expected that the animals with lesions of the MHb would not perceive the aversive effects, and therefore, would drink more of the nicotine-paired flavor than would the sham-operated animals.  In fact, on days 7-8 of conditioning, the lesioned animals did drink more of the nicotine-paired flavor than did the sham-operated animals, although they did not drink more of the saline-paired flavor.  Although other findings were less conclusive, the data support the idea that the MHb is, at least in part, responsible for mediating the aversive effects of nicotine.  More research should be done to further understand the role that the MHb plays in nicotine addiction, as this would help in the development treatments that may limit the intake of nicotine.

A Volumetric Analysis of the Amygdala as Related to Emotional Intelligence

Allison Kamm  (Psychology)

Advisor: Nancy Koven

As humans, we are constantly processing emotions, our own and others’ emotions. However, some do this better than others. The term ‘alexithymia’ has been given to individuals at the low end of this distribution and is marked by deficits in five areas 1) awareness of one’s emotions, 2) communication of emotions, 3) emotional display, 4) perception of others’ emotions and 5) abstract thought. While several functional MRI (fMRI) studies have linked decreased amygdala activation in response to emotional stimuli in alexithymics, to date no research has been done on potential structural abnormalities of the amygdala underlying this phenomenon. To determine possible correlations between amygdala volume and trait meta-mood components (ability to attend to emotion, to have clarity of emotion, and to repair emotions), I performed a manual volumetry analysis in 30 healthy adults in order to correlate trait meta-mood components, as measured by the Trait Meta-Mood Scale (TMMS), with unilateral and bilateral amygdala volume. Data including both genders yielded no correlations. However, negative correlations were found between right and bilateral amygdala volumes and the TMMS subscales of Clarity of Emotion and Mood Repair in female participants. This indicates that in general females with higher emotional processing have a reduced amygdala volume in comparison to females low in emotional processing skills. Additionally, no correlations were found in male participants.

Neuropeptide Phenylalanine (NPF) Induced Modulation of Phase 1 Feeding Behavior through Excitation of Buccal A Cluster (BAC) Neurons in The Pond Snail, Helisoma trivolvis

Kara Leasure (Neuroscience)

Advisor: Nancy Kleckner

In the pond snail, Helisoma trivolvis, three phases of feeding have been characterized: protraction (phase 1), retraction (phase 2), and hyperretraction (phase 3). Groups of interneurons and motor neurons make up the components of the central pattern generator (CPG) and are responsible for each phase. The phase 2 subunit is considered to be the controlling component of the CPG that drives the feeding pattern (Kleckner, unpublished). BAC neurons are a population of buccal neurons that can modulate and modify components of the CPG. BAC neurons can fire in response to sensory input and influence phase 1 and phase 3 motor neurons, resulting in behaviors such as regurgitation (protraction and retraction only) or satiation (cessation of feeding) (Arnett, 1996; Ramakrishnan, 2005; Sousa, 2008). Previous research indicates that Helisoma neuropeptide phenylalanine (NPF) activated BAC neurons may influence phase 1 and phase 3 motor neurons to produce satiation through a transition from regurgitation. To better understand how BAC neurons modify feeding and how BAC neuron activity effects phase 1 and phase 3 motor neurons, standard intracellular recording was used to study the simultaneous response of BAC neurons and phase 1 and phase 3 motor neurons to neuromodulators dopamine (DA) and NPF. BAC neuron and phase 1 motor neuron morphology was visualized with the injection of fluorescent tracers and imaged under epifluorescence microscopy. S1 motor neuron types (B6, B7, B8) and BAC neuron types (1, 2, 5, 6, 10, 13, 15, 16, 17, and novel) were identified using epifluorescent imaging. In saline, BAC neurons presented long, slow excitatory postsynaptic potentials (EPSP) in phase 2, phase 1 motor neurons showed characteristic firing in phase 1 followed by inhibitory postsynaptic potentials (IPSP) in phase 2, and phase 3 motor neuron B19 displayed phase 2 IPSPs with firing in phase 3. Results indicated DA increased the burst and action potential rates of S1 motor neurons and increased the burst rate of B19. BAC neuron slow EPSP rate increased predominantly and action potentials were inconsistently observed that corresponded with phase 2 IPSPs in phase 1 and phase 3 motor neurons. In NPF, phase 3 motor neurons were inhibited when BAC neurons fired action potentials, which was consistent with patterned regurgitation (Arnett, 1996; Sousa, 2008). NPF seemed to prolong activity in B6 and inhibit B7 and B8, thus it is possible that these results indicate a transition to satiation via regurgitation. BAC neurons were activated by NPF (types 1, 10, 16, and novel); this activation was also inconsistent but it did often inhibit phase 1 and phase 3 motor neuron activity. More research is needed to produce larger sample sizes of BAC neuron and phase 1 motor neuron types and to evaluate whether NPF induced BAC neuron activity induces satiation through regurgitation by interacting with phase 1 and phase 3 motor neurons.

The effects of small cardioactive peptide B (SCPB) on phase 3 motor neurons in the feeding cycle in Helisoma trivolvis

Olivia Moody (Neuroscience)

Advisor: Nancy Kleckner

In the pond snail, Helisoma trivolvis, a triphasic feeding system is controlled by a central pattern generator (CPG) in the buccal ganglia in the brain. The three phases normally fires in the order S1-S2-S3, which corresponds to the protraction of a tongue-like radula, retraction of the radula, and hyperretraction of the radula (Murphy, 2001). Small cardioactive peptide B (SCPB) is a neuropeptide thought to modulate the third phase (S3) of feeding, mimicking the excitatory effects of serotonin on S3 motor neurons (Murphy, Lukowiak & Stell, 1985). Using standard intracellular recording, the effects on the membrane potential of applying SCPB (1μM) and serotonin (1-3μM) on the buccal ganglia were measured from B19 (an S3 motor neuron) and compared.  Results indicate that SCPb and serotonin stimulates increased action potential frequency and burst rate, as measured in B19. Occasionally a slight hyperpolarization in the cell’s baseline membrane potential was seen in response to SCPB and a depolarization seen during serotonin application. This suggests that SCP­b may stimulate firing in B19 by a mechanism different than that by which serotonin stimulates firing. Furthermore, a novel inhibitory response to SCPB was seen in a cell thought to be the phase 3 motor neuron, B18. Understanding the role of neuropeptides in this feeding circuit allows us to better understand how a single flexible neuronal circuit can produce a variety of behaviors through chemical modulation.

The Effect of a Selective GABAA α5 Inverse Agonist on a Rat Model of PCP-Induced Schizophrenia

Lucy Cameron O’Keefe (Psychology)

Advisor: John Kelsey

The underlying neurophysiology of schizophrenia is still widely unknown. Research has shown the relevance of excess dopamine transmission through the effectiveness of typical antipsychotics, which block dopamine D2 receptors. However, these drugs only treat the positive symptoms of schizophrenia and have undesirable side effects. Recent research has implicated the activation of the GABAA α5 receptors in decreasing dopamine and positive-like symptoms. In this study, I examined the prediction that a GABAA α5 inverse agonist, MRK-016, should increase phencyclidine (PCP)-induced schizophrenia-like symptoms in rats in a hole board apparatus. However, in Experiment 1, 3 mg/kg MRK-016 actually reduced the PCP-induced increase in locomotion (a putative positive symptom), potentially reflecting a therapeutic effect, whereas in Experiment 2, 6 mg/kg MRK-016 tended to increase the effects of PCP on locomotion (a pro-psychotic effect), while decreasing the ability of PCP to decrease head dips (a putative negative symptom), reflecting an anti-psychotic effect. Thus my data offer only minimal support for the hypothesis that GABAA α5 agonists are likely to be antipsychotic.

Individual Differences that Moderate Brain Mechanisms of Anger

Helen Yuan (Neuroscience)

Advisor: Nancy Koven

When studying emotion, two competing models have prevailed: the valence/arousal model, which emphasizes the importance of valence (i.e., how positive and negative a feeling state is) and arousal (i.e., the intensity level of the feeling state), and the single-factor motivational model of affect, which argues that all emotions can be categorized by the single dimension of motivational bias. Neuroimaging has shown frontal lobe asymmetry to be critical for emotional valence, with left > right and right > left activity in the experience of positive and negative emotions, respectively. On the other hand, the motivational model predicts left > right and right > left frontal lobe activity for approach and avoidance behavioral tendencies, respectively. Anger, as a discrete emotion, pits these two models against each other, but little research has examined trait anger in the context of individual differences. In an attempt to relate frontal lobe asymmetry, anger, gender, and cultural background, six neuropsychological tests will be conducted to evaluate frontal asymmetry. The State Trait Anger Expression Inventory-2 (STAXI-2) will be used to measure trait anger, and the Individualism-Collectivism Interpersonal Assessment Inventory and the Self-Construal Scale will be used to assess cultural background. It is hypothesized that women and men will endorse internalizing and externalizing anger traits, respectively, while individuals from collectivistic and individualistic societies will endorse internalizing and externalizing anger traits, respectively. I further anticipate that individuals most prone to internalize anger will score better on neuropsychological tests sensitive to right frontal lobe functioning, whereas individuals most prone to externalize anger will score better on those sensitive to left frontal lobe functioning. Assuming these relationships to hold, mediation analyses will be conducted to determine if frontal lobe asymmetry mediates the relationships between gender/culture and anger profiles.