A choroid plexus apocrine secretion mechanism shapes CSF proteome and embryonic brain development.

We discovered that apocrine secretion by embryonic choroid plexus (ChP) epithelial cells contributes to the cerebrospinal fluid (CSF) proteome and influences brain development in mice. The apocrine response relies on sustained intracellular calcium signaling and calpain-mediated cytoskeletal remodeling. It rapidly alters the embryonic CSF proteome, activating neural progenitors lining the brain's ventricles. Supraphysiological apocrine secretion induced during mouse development by maternal administration of a serotonergic 5HT2C receptor agonist dysregulates offspring cerebral cortical development, alters the fate of CSF-contacting neural progenitors, and ultimately changes adult social behaviors. Critically, exposure to maternal illness or to the psychedelic drug LSD during pregnancy also overactivates the ChP, inducing excessive secretion. Collectively, our findings demonstrate a new mechanism by which maternal exposure to diverse stressors disrupts in utero brain development.

A brainstem circuit for phonation and volume control in mice.

Mammalian vocalizations are critical for communication and are produced through the process of phonation, in which expiratory muscles force air through the tensed vocal folds of the larynx, which vibrate to produce sound. Despite the importance of phonation, the motor circuits in the brain that control it remain poorly understood. In this study, we identified a subpopulation of ~160 neuropeptide precursor Nts (neurotensin)-expressing neurons in the mouse brainstem nucleus retroambiguus (RAm) that are robustly activated during both neonatal isolation cries and adult social vocalizations. The activity of these neurons is necessary and sufficient for vocalization and bidirectionally controls sound volume. RAm Nts neurons project to all brainstem and spinal cord motor centers involved in phonation and activate laryngeal and expiratory muscles essential for phonation and volume control. Thus, RAm Nts neurons form the core of a brain circuit for making sound and controlling its volume, which are two foundations of vocal communication.

Defining diurnal fluctuations in mouse choroid plexus and CSF at high molecular, spatial, and temporal resolution.

Transmission and secretion of signals via the choroid plexus (ChP) brain barrier can modulate brain states via regulation of cerebrospinal fluid (CSF) composition. Here, we developed a platform to analyze diurnal variations in male mouse ChP and CSF. Ribosome profiling of ChP epithelial cells revealed diurnal translatome differences in metabolic machinery, secreted proteins, and barrier components. Using ChP and CSF metabolomics and blood-CSF barrier analyses, we observed diurnal changes in metabolites and cellular junctions. We then focused on transthyretin (TTR), a diurnally regulated thyroid hormone chaperone secreted by the ChP. Diurnal variation in ChP TTR depended on Bmal1 clock gene expression. We achieved real-time tracking of CSF-TTR in awake TtrmNeonGreen mice via multi-day intracerebroventricular fiber photometry. Diurnal changes in ChP and CSF TTR levels correlated with CSF thyroid hormone levels. These datasets highlight an integrated platform for investigating diurnal control of brain states by the ChP and CSF.

Sex differences in long-term behavioral alterations, especially anxiety, following prenatal fluoxetine exposure in C57BL/6 mice.

Evidence demonstrates that psychiatric disorders during pregnancy are detrimental to the offspring. Many disorders are treated with SSRIs and increasing numbers of pregnant women now receive these drugs during gestation. The long-term neurobehavioral consequences of prenatal SSRI exposure require further evaluation. This study examined the effects of prenatal fluoxetine exposure in mice in an extensive battery of behaviors related to neurodevelopment, mood, social, and repetitive behaviors. C57BL/6J dams were administered fluoxetine at a low (0.6 mg/kg/day) or high (6 mg/kg/day) dose or saline from embryonic days 8 to 18. Juvenile mice were tested for changes in ultrasonic vocalizations and neuromotor development. In adulthood, offspring were tested for changes in behaviors related to anxiety, depression, social, and repetitive behaviors. Prenatal exposure to fluoxetine impaired surface righting reflex at P5, and sex-dependently reduced the frequency of ultrasonic vocalizations in juvenile males but not females. In adulthood, both males and females prenatally exposed to high, but not low, doses of fluoxetine exhibited an increase in repetitive behaviors in the marble burying task and a decrease in sucrose preference. Males, but not females, exposed to fluoxetine exhibited increased anxiety-related behaviors in the elevated plus maze. Prenatal fluoxetine exposure did not affect other adult behaviors including social preference, self-grooming, passive avoidance and open field activity. These findings suggest males are more sensitive than females to disruptions in serotonin balance during prenatal development and highlight the need for additional systematic and mechanistic studies to evaluate the impact of fluoxetine exposure during other periods of gestation.

A cellular and spatial map of the choroid plexus across brain ventricles and ages.

The choroid plexus (ChP) in each brain ventricle produces cerebrospinal fluid (CSF) and forms the blood-CSF barrier. Here, we construct a single-cell and spatial atlas of each ChP in the developing, adult, and aged mouse brain. We delineate diverse cell types, subtypes, cell states, and expression programs in epithelial and mesenchymal cells across ages and ventricles. In the developing ChP, we predict a common progenitor pool for epithelial and neuronal cells, validated by lineage tracing. Epithelial and fibroblast cells show regionalized expression by ventricle, starting at embryonic stages and persisting with age, with a dramatic transcriptional shift with maturation, and a smaller shift in each aged cell type. With aging, epithelial cells upregulate host-defense programs, and resident macrophages upregulate interleukin-1ß (IL-1ß) signaling genes. Our atlas reveals cellular diversity, architecture and signaling across ventricles during development, maturation, and aging of the ChP-brain barrier.

Tracking Calcium Dynamics and Immune Surveillance at the Choroid Plexus Blood-Cerebrospinal Fluid Interface.

The choroid plexus (ChP) epithelium is a source of secreted signaling factors in cerebrospinal fluid (CSF) and a key barrier between blood and brain. Here, we develop imaging tools to interrogate these functions in adult lateral ventricle ChP in whole-mount explants and in awake mice. By imaging epithelial cells in intact ChP explants, we observed calcium activity and secretory events that increased in frequency following delivery of serotonergic agonists. Using chronic two-photon imaging in awake mice, we observed spontaneous subcellular calcium events as well as strong agonist-evoked calcium activation and cytoplasmic secretion into CSF. Three-dimensional imaging of motility and mobility of multiple types of ChP immune cells at baseline and following immune challenge or focal injury revealed a range of surveillance and defensive behaviors. Together, these tools should help illuminate the diverse functions of this understudied body-brain interface.

A concerted metabolic shift in early forebrain alters the CSF proteome and depends on MYC downregulation for mitochondrial maturation.

Massive, coordinated cellular changes accompany the transition of central nervous system (CNS) progenitors from forebrain neurectodermal cells to specified neuroepithelial cells. We have previously found that MYC regulates the changing ribosomal and proteostatic landscapes in mouse forebrain precursors at embryonic days E8.5 and E10.5 (before and after neural tube closure; NTC) (Chau et al., 2018). Here, we demonstrate parallel coordinated transcriptional changes in metabolic machinery during this same stage of forebrain specification. Progenitors showed striking mitochondrial structural changes transitioning from glycolytic cristae at E8.5, to more traditional mitochondria at E10.5. Accordingly, glucose use shifted in progenitors such that E8.5 progenitors relied on glycolysis, and after NTC increasingly used oxidative phosphorylation. This metabolic shift was matched by changes in surrounding amniotic and cerebrospinal fluid proteomes. Importantly, these mitochondrial morphological shifts depend on MYC downregulation. Together, our findings demonstrate that metabolic shifting accompanies dynamic organelle and proteostatic remodeling of progenitor cells during the earliest stages of forebrain development.

Peripheral Mechanosensory Neuron Dysfunction Underlies Tactile and Behavioral Deficits in Mouse Models of ASDs.

Patients with autism spectrum disorders (ASDs) commonly experience aberrant tactile sensitivity, yet the neural alterations underlying somatosensory dysfunction and the extent to which tactile deficits contribute to ASD characteristics are unknown. We report that mice harboring mutations in Mecp2, Gabrb3, Shank3, and Fmr1 genes associated with ASDs in humans exhibit altered tactile discrimination and hypersensitivity to gentle touch. Deletion of Mecp2 or Gabrb3 in peripheral somatosensory neurons causes mechanosensory dysfunction through loss of GABAA receptor-mediated presynaptic inhibition of inputs to the CNS. Remarkably, tactile defects resulting from Mecp2 or Gabrb3 deletion in somatosensory neurons during development, but not in adulthood, cause social interaction deficits and anxiety-like behavior. Restoring Mecp2 expression exclusively in the somatosensory neurons of Mecp2-null mice rescues tactile sensitivity, anxiety-like behavior, and social interaction deficits, but not lethality, memory, or motor deficits. Thus, mechanosensory processing defects contribute to anxiety-like behavior and social interaction deficits in ASD mouse models. PAPERCLIP.