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Principal Investigator: Daniel Charles Castro
Organization: WASHINGTON UNIVERSITY
Fiscal Year: 2024
Award: $523,855
Funding agency: National Institute of Mental Health
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.
Terms: <21+ years old><Active Follow-up><Adult><Adult Human><Affect><Affective><Analgesic Agents><Analgesic Drugs><Analgesic Preparation><Analgesics><Anatomic Sites><Anatomic structures><Anatomy><Anodynes><Antinociceptive Agents><Antinociceptive Drugs><Appetite><Appetite stimulated><Behavior><Behavioral><Biological><Bite><Brain><Brain Nervous System><CRISPR><CRISPR approach><CRISPR based approach><CRISPR method><CRISPR methodology><CRISPR technique><CRISPR technology><CRISPR tools><CRISPR-CAS-9><CRISPR-based method><CRISPR-based technique><CRISPR-based technology><CRISPR-based tool><CRISPR/CAS approach><CRISPR/Cas method><CRISPR/Cas system><CRISPR/Cas technology><CRISPR/Cas9><CRISPR/Cas9 technology><Calcium><Cas nuclease technology><Cell Communication and Signaling><Cell Nucleus><Cell Signaling><Characteristics><Chronic><Clustered Regularly Interspaced Short Palindromic Repeats><Clustered Regularly Interspaced Short Palindromic Repeats approach><Clustered Regularly Interspaced Short Palindromic Repeats method><Clustered Regularly Interspaced Short Palindromic Repeats methodology><Clustered Regularly Interspaced Short Palindromic Repeats technique><Clustered Regularly Interspaced Short Palindromic Repeats technology><Code><Coding System><Color><Data><Desire for food><Devices><Dissociation><Dorsal><E-stim><Eating><Electric Stimulation><Encephalon><Endoscopy><Enkephalin Receptors><Enkephalins><FISH Technic><FISH Technique><FISH analysis><FISH assay><Fluorescence In Situ Hybridization><Fluorescent in Situ Hybridization><Food Intake><Genetic><Goals><Health Care Costs><Health Costs><Healthcare Costs><Image><Increased food appetite><Individual><Infumorph><Intracellular Communication and Signaling><Kadian><Lateral><Ligands><MS Contin><MSir><Measures><Mediating><Mental disorders><Mental health disorders><Mesencephalic Central Gray><Mesencephalon><Mice><Mice Mammals><Microinjections><Mid-brain><Midbrain><Midbrain Central Gray><Midbrain structure><Moods><Morphia><Morphine><Motivation><Murine><Mus><NIMH><National Institute of Mental Health><Nerve Cells><Nerve Unit><Neural Cell><Neurocyte><Neurons><Nucleus><Nucleus Accumbens><Opiate Peptides><Opiate Receptors><Opiate agonist><Opiate receptor agonist><Opiates><Opioid><Opioid Peptide><Opioid Receptor><Opioid agonist><Opioid receptor agonist><Opsin><Oramorph><Oramorph SR><Pattern><Peptides><Periaqueductal Gray><Pharmacology><Phenotype><Photons><Pilot Projects><Play><Process><Psychiatric Disease><Psychiatric Disorder><Receptor Protein><Regulation><Rewards><Rod-Opsin><Role><Roxanol><Signal Transduction><Signal Transduction Systems><Signaling><Site><Statex SR><Strategic Planning><System><Tail><Technology><Testing><Therapeutic Intervention><United States><Virus><active followup><adulthood><annulus of the aqueduct><antagonism><antagonist><behavior test><behavioral test><biologic><biological adaptation to stress><biological signal transduction><calcium indicator><cell type><delta opioid receptor><design><designing><dorsal raphe nucleus><electrostimulation><endogenous opiate><endogenous opioids><endoscopic imaging><experience><experiment><experimental research><experimental study><experiments><fluorophore><follow up><follow-up><followed up><followup><imaging><imaging in vivo><in vivo imaging><increased appetite><increased hunger><insight><intervention therapy><knock-down><knockdown><mental illness><midbrain central gray substance><motivated behavior><mu receptors><neural><neural circuit><neural circuitry><neural control><neural regulation><neurobiological mechanism><neurochemical><neurochemistry><neurocircuitry><neuromodulation><neuromodulatory><neuronal><neuropsychiatric><neuropsychiatric disease><neuropsychiatric disorder><neuropsychiatry><neuroregulation><next generation><optogenetics><pain killer><pain medication><pain reliever><painkiller><periaqueductal gray matter><pharmacologic><pilot study><psychiatric illness><psychological disorder><reaction; crisis><receptor><receptor expression><redshift><response><social role><stress response><stress; reaction><synaptic circuit><synaptic circuitry><targeted drug therapy><targeted drug treatments><targeted therapeutic><targeted therapeutic agents><targeted therapy><targeted treatment><wireless><wireless implant><δ OR><δ ORs><δ opioid receptors><δ-OR><δ-ORs><δOR><δORs><μ receptors>