Abstract

Chemogenetic and Optogenetic Manipulation of Neuronal Projections from the Lateral Hypothalamus to the Paraventricular Nucleus of the Thalamus Impact Cue-Motivated Behaviors

Iglesias A, Westbrook S, Chang SE, Chung, J, Campus P, Flagel SB
61st Annual Meeting of the American College of Neuropsychopharmacology. 2022.

Background: The survival of an organism is dependent on their ability to properly respond to cues in the environment. Associative learning processes underlie an individual’s response to such cues. For some, reward-paired cues are attributed with incentive motivational value (incentive salience), and can gain excessive control, leading to maladaptive behavior that is characteristic of psychopathology. To investigate the neural mechanisms that promote incentive learning, we utilize the goal-tracker (GT)/ sign-tracker (ST) animal model. With Pavlovian conditioned approach (PavCA) training, GTs attribute predictive value to reward-cues; whereas STs attribute predictive and incentive value to such cues. The attribution of incentive motivational value, or incentive salience, transforms the cue into an attractive and desirable stimulus. For STs, both food- and drug-associated cues gain excessive incentive value and elicit maladaptive behaviors. The GT/ST model, therefore, can be utilized to elucidate the neurobiological mechanisms that encode adaptive or maladaptive cue-driven behaviors. GTs and STs rely on distinct neurobiological mechanisms and the paraventricular nucleus of the thalamus (PVT) has emerged as a neural hub that mediates their characteristic differences in associative learning. Prior findings have led us to postulate that bottom-up projections to the PVT relay the incentive value of reward-associated cues, with the lateral hypothalamus (LH), which sends dense orexinergic (OXergic) projections to the PVT, acting as a critical neural node. Orexin (OX) is known to play a role in motivation, and administration of OX receptor antagonists into the PVT attenuates the incentive value of food-paired cues in STs. We hypothesize that OXergic transmission in the LH-PVT pathway encodes the incentive motivational value of reward cues and modulates the propensity to sign-track. Thus, we postulate that excitation of the pathway, via optogenetics, would elicit sign-tracking, and inhibition of the pathway following learning, via chemogenetics, would reduce sign-tracking behavior.

Methods: Here we investigate the role of the LH-PVT pathway in encoding the value of reward cues via (1) optogenetic excitation and (2) chemogenetic inhibition during PavCA behavior. In PavCA sessions, an illuminated cue (lever) extends for 8 seconds then retracts and a food pellet is dispensed into a food cup. In the optogenetics study, we used transgenic OX-Cre Long Evans male and female rats (N = 11), which express Cre-recombinase in OX neurons, and infused a retrograde Cre-dependent excitatory optogenetic virus (pAAVrg-Ef1α-DIO-ChR2-EYFP; channelrhodopsin, ChR2) or empty vector (pAAVrg-Ef1α-DIO-EYFP; control) into the anterior PVT. Animals received laser-induced excitation of LH-PVT neurons during cue (lever) presentation on sessions 1-5.

In the chemogenetic study, we utilized a dual-vector approach to selectively express an inhibitory (Gi) DREADD virus in the LH-PVT pathway in outbred Sprague-Dawley male rats (N = 46). A retrograde Cre virus (pAAVrg-hSyn-EGFP-Cre) was infused into the anterior and posterior PVT, while a Cre-dependent Gi virus (pAAV8-hSyn-DIO-hM4D-mCherry) was bilaterally infused into the LH. Following virus incubation, rats had 7 sessions of PavCA. After acquiring a conditioned response, rats received either vehicle or clozapine-N-oxide (CNO; 5 mg/kg) prior to sessions 5-7 of PavCA to activate the Gi-DREADDs.

Results: In the optogenetics study, preliminary results indicate that OXergic LH-PVT pathway stimulation during cue presentation results in increased goal-tracking behavior in ChR2 (n = 6) rats compared to controls (n = 5). This effect appears to be more pronounced in male ChR2 (n = 3) rats relative to male EYFP (n = 2), female ChR2 (n = 3), and female EYFP (n = 3); however, studies are ongoing to increase the sample size.

In the chemogenetics study, we assessed the impact of Gi DREADD activation on the expression of a learned conditioned response, session 5-7 of PavCA. A linear mixed-effects model analysis compared the effect of treatment (VEH vs. CNO) and phenotype (ST vs GT vs IR (intermediate responders)) for various behaviors in PavCA across sessions, and for all measures, there was a significant effect of phenotype (P < 0.005). When phenotypes were assessed independently using a repeated measures ANOVA, there was no effect of treatment on sessions 5-7 for either STs (VEH, n = 3; CNO, n = 5) or IRs (VEH, n = 7; CNO, n = 7). However, for GTs, there was a significant effect of treatment, such that CNO-treated GTs (n = 8) had less food-cup contacts (p = 0.034), a lower probability to approach the food-cup (p = 0.002), and a slower food-cup approach (p = 0.01) than VEH-treated GTs (n = 16).

Conclusions: These findings suggest that the LH-PVT pathway plays an important role in goal-directed behaviors. Preliminary findings indicate that optogenetic stimulation of OXergic neurons in the LH-PVT pathway of transgenic male rats increases goal-tracking behavior. Further, chemogenetic inhibition of the LH-PVT pathway in outbred male rats decreases the expression of goal-directed behaviors for GTs, without affecting the behavior of STs or IRs. Thus, the LH-PVT pathway appears to modulate predictive value encoding and the expression of goal-directed behaviors both early in training and after a conditioned response has been acquired. Ongoing studies are increasing the sample size in both studies to further investigate a potential role for the LH-PVT pathway in incentive learning and any sex differences that might arise.