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1. Mechanisms of D2R signaling in the blood brain barrier that regulates courtship in Drosophila melanogaster

   Gautam, Sumit.; Love, Cameron R.; Dauwalder, Brigitte (March 2023, Drosophila Research Conference)

   Abstract Flies have an open circulatory system and their Blood Brain Barrier (BBB) surrounds the brain like a tight cap. The fly BBB consists of two layers of glial cells. The outer layer is formed by the Perineurial Glia (PG). The inner BBB layer consists of the Subperineurial Glia (SPG) that form the tight barrier that, like its mammalian counterpart, acts both as a diffusion and a xenobiotic transport barrier. Underneath the SPG lie the neuronal cell bodies. The Drosophila BBB shows the same barrier properties as the mammalian barrier and profiling of Drosophila BBB cells has shown a high degree of molecular conservation. We have previously shown that the Drosophila BBB plays a sex-specific role in regulating behavior. Conditional adult feminization of SPG cells in otherwise normal males leads to significantly reduced courtship. In agreement with this, in a microarray screen of isolated SPG cells, we identified a number of male-enriched transcripts. One of them encodes the dopamine-2 like receptor (D2R). We have found that conditional knockdown of D2R in adult male Drosophila SPG decreases courtship. Likewise, D2R mutant males have courtship defects. They can be rescued by expression of wildtype D2R in the SPG cells of mature adult males, demonstrating a physiological requirement for the receptor in these cells for courtship control4. The D2R receptor is highly conserved. It has been found that it can act via biased signaling (through G protein or b-arrestin) in mammals. We have previously found that signaling through Gao and arrestin in the BBB is required for proper male courtship. Although D2R is best known for signaling through cAMP, we have not found a requirement for cAMP/PKA signaling for courtship. To investigate the signaling pathways downstream of D2R that are responsible for courtship control we have mutagenized D2R proteins and examined their ability to rescue D2R mutants. Based on Peterson et al. we designed proteins capable of G-protein or arrestin biased signaling, respectively, and tested their ability to rescue the courtship defects of D2R mutants. Our data suggest that D2R signaling through b-arrestin is a major mediator of BBB courtship control. 

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2. Ecdysone In The Blood-Brain Barrier And Male Courtship Behavior

   Waqar, M.; Dauwalder, B. (March 2023, Drosophila Research Conference)

   Abstract: We have previously found that the physiology of the blood-brain barrier (BBB) plays an important role in the regulation of male courtship behavior. Recently, we have shown that the nuclear hormone receptor Hr46 (Hormone receptor-like in 46) is required in the BBB of adult males for normal courtship in Drosophila melanogaster (Lama et al. 2022). Hr46 is well-characterized in development, where it is induced by the hormone Ecdysone and is a major mediator of the Ecdysone response. Using a reporter construct, we have found that Ecdysone is present in the BBB; however, nothing is known about its potential roles in these cells. Since, Hr46 is a major mediator of Ecdysone signaling; we hypothesize that Ecdysone is required in the BBB in the regulation of male courtship behavior. To test our hypothesis, we are using several different strategies to examine the role of Ecdysone and Ecdysone effectors in the BBB for courtship. These include the knockdown of the Ecdysone receptor (EcR) in mature males and lowering the levels of Ecdysone in the BBB by overexpressing an Ecdysone degrading enzyme. We are also testing the role of alternative Ecdysone effectors like DopEcR and an Ecdysone transporter in the BBB. 

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3. Feminization of the Blood–Brain Barrier Changes the Brain Transcriptome of Drosophila melanogaster Males

   Davis, Danyel S; Hashem, Warda; Lama, Chamala; Reeve, Joseph L; Dauwalder, Brigitte (August 2025, Curr Issues Mol Biol)

   Beyond its crucial role as a tight barrier to protect the nervous system, the Blood–Brain Barrier (BBB) is increasingly being recognized for its physiological processes that affect brain function and behavior. In Drosophila melanogaster, the BBB expresses sex-specific transcripts, and a change in the sexual identity of adult BBB cells results in a significant reduction in male courtship behavior. The molecular nature of this BBB/brain interaction and the molecules that mediate it are unknown. Here we feminize BBB cells by targeted expression of the Drosophila female-specific master regulator TraF in otherwise normal males. We examined the effect on RNA expression in dissected brains by RNA sequencing. We find that 283 transcripts change in comparison to normal control males. Transcripts representing cell signaling processes and synaptic communication are enriched, as are hormonal mediators. These transcripts provide a valuable resource for addressing questions about BBB and brain interaction. 

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4. A Soluble Platelet-Derived Growth Factor Receptor-β Originates via Pre-mRNA Splicing in the Healthy Brain and Is Upregulated during Hypoxia and Aging

   https://doi.org/10.3390/biom13040711  

   Payne, Laura Beth; Abdelazim, Hanaa; Hoque, Maruf; Barnes, Audra; Mironovova, Zuzana; Willi, Caroline E.; Darden, Jordan; Houk, Clifton; Sedovy, Meghan W.; Johnstone, Scott R.; et al (April 2023, Biomolecules)

   The platelet-derived growth factor-BB (PDGF-BB) pathway provides critical regulation of cerebrovascular pericytes, orchestrating their investment and retention within the brain microcirculation. Dysregulated PDGF Receptor-beta (PDGFRβ) signaling can lead to pericyte defects that compromise blood-brain barrier (BBB) integrity and cerebral perfusion, impairing neuronal activity and viability, which fuels cognitive and memory deficits. Receptor tyrosine kinases such as PDGF-BB and vascular endothelial growth factor-A (VEGF-A) are often modulated by soluble isoforms of cognate receptors that establish signaling activity within a physiological range. Soluble PDGFRβ (sPDGFRβ) isoforms have been reported to form by enzymatic cleavage from cerebrovascular mural cells, and pericytes in particular, largely under pathological conditions. However, pre-mRNA alternative splicing has not been widely explored as a possible mechanism for generating sPDGFRβ variants, and specifically during tissue homeostasis. Here, we found sPDGFRβ protein in the murine brain and other tissues under normal, physiological conditions. Utilizing brain samples for follow-on analysis, we identified mRNA sequences corresponding to sPDGFRβ isoforms, which facilitated construction of predicted protein structures and related amino acid sequences. Human cell lines yielded comparable sequences and protein model predictions. Retention of ligand binding capacity was confirmed for sPDGFRβ by co-immunoprecipitation. Visualizing fluorescently labeled sPDGFRβ transcripts revealed a spatial distribution corresponding to murine brain pericytes alongside cerebrovascular endothelium. Soluble PDGFRβ protein was detected throughout the brain parenchyma in distinct regions, such as along the lateral ventricles, with signals also found more broadly adjacent to cerebral microvessels consistent with pericyte labeling. To better understand how sPDGFRβ variants might be regulated, we found elevated transcript and protein levels in the murine brain with age, and acute hypoxia increased sPDGFRβ variant transcripts in a cell-based model of intact vessels. Our findings indicate that soluble isoforms of PDGFRβ likely arise from pre-mRNA alternative splicing, in addition to enzymatic cleavage mechanisms, and these variants exist under normal physiological conditions. Follow-on studies will be needed to establish potential roles for sPDGFRβ in regulating PDGF-BB signaling to maintain pericyte quiescence, BBB integrity, and cerebral perfusion—critical processes underlying neuronal health and function, and in turn, memory and cognition. 

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5. GPR37 Regulates Granule Cell Maturation and Affective Behavior in the Hippocampus via Sex-Dependent Mechanisms (Abstract ID: 275646)

   https://doi.org/10.1016/j.jpet.2026.104807  

   Mondaizie, Kaylor; Owino, Sharon; Fernandez, Franchesa (May 2026, The Journal of Pharmacology and Experimental Therapeutics)

   Hippocampal neurogenesis is a lifelong form of neural plasticity in which newly generated granule neurons mature and integrate into circuits supporting cognition and emotional regulation. In the dentate gyrus, these neurons are continuously produced throughout adulthood and contribute to region-specific functions along the dorsoventral axis. Although adult neurogenesis proceeds through tightly regulated stages, the mechanisms governing late-stage granule cell maturation—particularly those involving canonical Wnt/β-catenin signaling via the co-receptor LRP6—remain unresolved. Emerging evidence suggests that Wnt signaling can be fine-tuned at the level of receptor and co-receptor interactions, with recent studies identifying the orphan G protein–coupled receptor GPR37 as a potential upstream modulator of Wnt signaling through its interaction with LRP6, where it may function as an endoplasmic reticulum chaperone to potentiate Wnt/β-catenin signaling. The direct functional role of GPR37 in adult hippocampal neurogenesis, however, has not been explored. Notably, late-stage maturation of adult-born granule cells in the ventral hippocampus—an area critical for emotional regulation—is strongly associated with anxiety-related behaviors. We hypothesize that the GPR37–LRP6–Wnt/β-catenin axis constitutes a novel regulatory network promoting late-stage granule cell maturation, and that disruption of this pathway alters anxiety-related behavior. Male and female wild-type and Gpr37⁻/⁻ mice (2–4 months) were used to assess hippocampal morphology and anxiety-related behaviors. Dentate gyrus cell populations were quantified using immunohistochemistry, RNAscope, and IMARIS to assess markers of immature and mature granule cells. Soma size and neuronal eccentricity were quantified using confocal imaging and CellProfiler. Anxiety-related behavior was evaluated using the light/dark box and elevated plus maze, with DeepLabCut providing high-resolution behavioral tracking. Analyses were sex-balanced, blinded, and conducted in RStudio and GraphPad Prism. Loss of GPR37 did not alter total nuclei or neuronal number in the dentate gyrus; however, female Gpr37⁻/⁻ mice exhibited increased neuronal eccentricity and larger soma size. Gpr37⁻/⁻ mice spent less time in the light compartment and produced more boli in the light/dark box, indicating increased anxiety-like behavior. Elevated plus maze analyses revealed sex-influenced behavioral differences, with Gpr37⁻/⁻ males spending more time in the center region compared to females. Together, these findings indicate that GPR37 positively regulates granule cell maturation and anxiety-related behavior, with emerging sex-dependent effects potentially mediated through LRP6-dependent Wnt/β-catenin signaling. Ongoing studies will further define direct GPR37–LRP6 regulation of adult neurogenesis and affective outcomes. 

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