Saturday, May 17, 2025
5/17/2025
Investigating non-canonical mechanisms of endogenous opioids on motivation in dorsal midbrain - Project Summary The primary goal of this R01 is to determine the computational and functional role of endogenous opioids in specific dorsal midbrain nuclei on motivated behaviors. The preponderance of mental illness in the United States results in tens of millions of dollars in healthcare costs. While many neuropsychiatric conditions can be dissociated based on the presence or absence of specific features, a common theme across mental illnesses is the dysregulation of affective or motivated behaviors. The endogenous opioid system is known to powerfully modulate affective and motivational neural circuits. Historically, dorsal midbrain nuclei (including ventrolateral periaqueductal gray nucleus and the adjacent dorsal raphe nucleus) have been shown to be important sites for opioid action. More recently, the lateral dorsal raphe nucleus subregion (LDRN) and nucleus accumbens were shown to be important sites in an opioid-mediated mesolimbic circuit of appetitive motivation. However, while downstream opioid activity in this LDRNaccumbens circuit specifically enhances appetitive motivation, convergent studies indicate that endogenous opioids may play a motivationally suppressive role within LDRN itself. For example, experiments in the 1980s demonstrated morphine microinjections into ventrolateral PAG/LDRN could suppress food intake. Correspondingly, pilot studies in our lab using a CRISPR-Cas9 mediated knockdown of opioid peptides oppositely facilitated food intake. Furthermore, local LDRN opioid activity appears to suppress both appetitive and aversive motivated behaviors (e.g., local opioid antagonism in LDRN increases defensive or escape behaviors). These broad, anti-motivational opioid effects suggest that dysregulation could affect a wide range of affective or motivated behaviors. Therefore, the goal of this R01 application is to determine how endogenous opioids regulate appetitive and aversive motivated behaviors in LDRN by multiplexing genetic, pharmacological, in vivo imaging, and optogenetic technologies. First, we will identify the anatomical characteristics of opioids within dorsal midbrain nuclei (Aim 1). In tandem we will test the functional localization of opioids by performing receptor selective pharmacological antagonism via wireless fluidic devices and CRISPR-Cas9 knockdown of opioid peptides. Next, we will use dual-color 1-photon endoscopic imaging to examine how local opioidergic and non-opioidergic neurons interact to encode appetitive and aversive behaviors (Aim 2). Follow up experiments will use simultaneous 1-photon imaging with cell-type selective optogenetic neuromodulation to augment or disrupt LDRN encoding and expression of motivated behaviors. Finally, we will multiplex 1-photon imaging, CRISPR-Cas9 knockdown, and cell-type specific optogenetic stimulation to determine how endogenous opioid peptides casually augment LDRN encoding and expression of motivated behaviors (Aim 3). Together, these studies may provide insights into how neuropsychiatric disorders are often characterized by affective and motivational dysregulation, and suggest specific neurochemical targets for therapeutic intervention.
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