Which specific brain projections are involved in natural reward?

The way the brain processes rewards is an important building block for animal behavior. In many mental disorders, the parts of the brain that deal with reward processing are not functioning normally. To find out more how this reward system and related structures works, we studied normal behavior that is rewarding, sex. The reward system in the brain is not a single center in the brain, but a collection of brain regions. Together, they are essential for an animal’s understanding when things are nice and when things are not. This in part directs behavior: when a situation or a behavior is pleasurable, we want to do it again, but we avoid repeating the situations and behaviors that were unpleasant. This mechanism makes animals hardwired to recognize behaviors and situations that are important for the survival of the animal or the species, such as feeding, drinking, social behaviors and sex. With this project, we tried to find out more about how specific parts of the reward system are connected and how they are involved in regulating sexual behavior, which is a naturally rewarding behavior. These brain regions are called the preoptic area (POA), medial amygdala (MeA) and bed nucleus of stria terminalis (BNST). To study these regions in rats, we wanted to see the fluctuations of activity levels of these brain regions during sexual behavior and we wanted to see if sexual behavior changed if we turned these regions on or off. To do so, we used fiber photometry to register activity levels, and chemogenetics to turn specific cells on or off. With these techniques, we found out that the medial amygdala (MeA) and the bed nucleus of the stria terminalis (BNST) did not react the same to all types of sexual behavior. The BNST became active earlier than the MeA when a male rat was mounting or intromitting, while these brain regions fired simultaneously when the animal ejaculated. We also found that these regions did not regulate the strength of the initial approach response of the male animal to a female. The BNST appears to determine how much time it takes for the male to ejaculate given the right circumstances. In another experiment, we looked at the preoptic area, a brain region that receives a lot of information from the MeA and BNST. The preoptic area becomes gradually more active before mounts and intromissions, and peaks shortly after. Before an ejaculation this brain region is especially active. These activity patterns appear to become more focused when the male becomes more sexually experienced. We concluded that this brain region plays a role in making the transition from copulation to ejaculation. Because the amygdala is so strongly connected to the preoptic area, we wanted to find out how this specific connection in isolation was involved in the regulation of sexual behavior. This time we did not only look inside the brain with fiber photometry, but we also manipulated the brain cells that connected the amygdala to the preoptic area. With chemogenetics we are able to turn those connections off during different stages of sexual experience. Looking at the behavior itself, and the activity in the brain, we can see if this intervention over time disrupts gaining of sexual experience. Finally, we tried to find out if the BNST was acting in the same way when the rat interacted with different natural rewards. For this, we compared between two things that rats enjoy, sex and sugar. We found out that the BNST seems to modulate the tempo and the intervals of the engagement the animals had with both sex and sugar. This suggests that different types of natural rewards trigger the same wiring in the brain to regulate motivated behaviors. This project has given us more insight into the role certain brain areas play in the regulation of sexual motivation and behavior. Developing our understanding of how the brain works when we interact with these kinds of rewards is necessary to figure out what might go wrong in the development of certain mental disorders.

The reward system in the brain is an essential element involved in motivation and responsible for many of our behaviors. A pleasant feeling that coincides with a certain behavior (pattern) makes us want to repeat that behavior, while we avoid those actions that had a less desirable outcome. This project has provided a fundamental understanding of how the preoptic area (POA), medial amygdala (MeA) and bed nucleus of the stria terminalis (BNST) were involved in regulating natural reward and how these brain regions communicate. Sexual behavior was used as a natural incentive-driven behavior to study the motivational system in rats. Sexual intercourse, or copulation, will never occur without approach behavior, and thus, without an intrinsic state of sexual motivation. Altogether, this project has, and still will, generate more insights into the role certain brain areas play regulating in sexual motivation and behavior. By understanding how our brain regulates this kind of rewarding behaviors, it becomes possible to study what can go wrong and lead to mental disorders. On the long-term, our project will thus potentially contribute to finding treatments for disorders linked to reward. One of the most impactful outcomes of the project is the development of a new scoring scheme for evaluating male rat sexual behavior in more detail. This advanced the behavioral assessment of sexual behavior by including an in-depth analysis of the temporal patterning of different sexual behaviors and the breaks between them. This temporal patterning and the intensity of behaviors, as well as the breaks between them, determine the efficiency of copulation, and can thereby generate a more complete understanding of the behavioral effects. The goal is that more researchers in the world will start using this new behavioral assessment tool when studying male rat sexual behavior. Moreover, besides the practical outcomes of 4 research articles (plus three more to come), 1 review article, 1 book chapter, and 20 outreach activities on scientific conferences and public events, the project has also led to increased international research collaborations that resulted in the development of new research ideas and grant proposals.

Disturbances in the reward system are related to many disorders like addiction, obesity, anhedonia, and sexual dysfunctions. The disadvantage of previous research is that the reward system is manipulated by unnatural external factors like cocaine or alcohol. This research project uses sexual behavior as a more natural incentive-driven behavior to study the motivational system in rats. Another advantage of studying sexual behavior is that both motivation and the transition to actual consummatory behavior can be studied separately in this model for natural reward. This project will investigate which pathways in the brain are involved in sexual motivation and behavior. The projections of interest are the projections from the medial amygdala (MeA) and the bed nucleus of the stria terminalis (BNST) to the preoptic area (POA). Interestingly, these brain areas play dual roles in the regulation of sexual behavior; inhibitory on motivation and stimulatory on copulation. This proposes that they should switch from an active state into an inactive state in order to fulfil all aspects of sexual behavior. This project will investigate the role of the specific projections between the MeA, BNST and POA in the regulation of sexual motivation and the transition to behavior. The project uses the very advanced and novel techniques that can specifically target crucial connections between brain regions (projections) in rats: fiber photometry and chemogenetics. Fiber photometry works by injecting adeno-associated viral (AAV) vectors in brain regions. These vectors carry DNA that encodes calcium indicators. Transfected cells in that region consequently express calcium indicators in their cell bodies, axons and in their downstream projections that synapse on neurons in other regions. Activation of neurons occurs with rapidly increasing levels of intracellular calcium. This calcium instantly binds to the calcium indicators in the cells, resulting in the emission of a fluorescent signal that can be detected by the implanted fiber. By implanting an optic fiber into a certain brain region, fiber photometry can measure the neural activity of this brain region or specific projection on a millisecond timescale. Chemogenetics also makes use of AAV vectors, which in this case are designed to express clozapine-N-oxide (CNO)-sensitive G-protein-coupled receptors (either stimulating or inhibiting neuronal activity) in neurons. By injecting CNO, chemogenetics enables us to turn on or off the specific projection. Activating or inhibiting specific projections using chemogenetics enables us to isolate their contributions to the emergent behavior. This project will thus generate more insights into the role of the communication between brain areas in sexual motivation and behavior. This can help finding treatments for disorders linked to reward.