(119a) Developing an Ex Vivo Mesolimbic Pathway Model for Exploring Phenotypes Linked with Addiction

The mesolimbic pathway, the connection between dopaminergic (DA) neurons of the midbrain and medium spiny neurons (MSNs) of the ventral forebrain, is of critical importance because it has been linked with reward behaviors such as addiction. Repeated exposures to drugs of abuse lead to long-term neuroplastic alterations, with ample evidence of molecular and cellular level changes in rodent models. To study these mechanisms in human backgrounds, researchers need a model system which can be used to manipulate and dynamically monitor both the short term dynamic electrical and molecular response and the long-term alterations produced by chronic drug use. Stem cell derived human cerebral organoids are good model candidates as they can be differentiated into specific neural subtypes that can mimic cellular responses seen in vivo and can be dynamically perturbed. Previous work has generated ventral forebrain and midbrain organoids, however they used different cell lines and medias to generate their organoids. Here we modified a whole brain organoid protocol to generate both DA+ midbrain and MSN+ ventral forebrain organoids while ensuring their media components are the same after 25 days of culture, allowing for organoid fusion. Cellular identity and functional activity were confirmed before exposing ventral forebrain organoids to acute and chronic dosing regiments of dopamine and ventral forebrain-midbrain organoid fusions to acute and chronic cocaine dosing. Through a combination of calcium imaging, and single cell sequencing we have produced a comprehensive characterization of this organoid system’s electrical and cell type specific transcriptional responses to acute and chronic stimuli. The results demonstrate cerebral organoids’ ability to recreate known phenotypes including increased expression of early genes FosB and cFos and pathway genes linked with increased synaptogenesis and complex phenotypes including crosstalk between drugs of abuse and molecular and cellular desensitization. We are advancing these models further by developing dynamic reporters of target genes and pathways to better understand underlying mechanisms of and provide platforms for screening potential therapeutics.