Hazel Sive

16p11.2 deletion syndrome is a severe genetic disorder associated with the deletion of 27 genes from a Copy Number Variant region on human chromosome 16. Symptoms associated include cognitive impairment, language and motor delay, epilepsy or seizures, psychiatric disorders, autism spectrum disorder (ASD), changes in head size and body weight, and dysmorphic features, with a crucial need to define genes and mechanisms responsible for symptomatology. In this review, we analyze the clinical associations and biological pathways of 16p11.2 locus genes and identify that a majority of 16p11.2 genes relate to metabolic processes. We present a hypothesis in which changes in the dosage of 16p11.2 metabolic genes contribute to pathology through direct or indirect alterations in pathways that include amino acids or proteins, DNA, RNA, catabolism, lipid, energy (carbohydrate). This hypothesis suggests that research into the specific roles of each metabolic gene will help identify useful therapeutic targets.

Collecting eggs from adultXenopus laevisandXenopus tropicalisto raise healthy embryos and tadpoles is relatively simple but requires careful handling of the frog. Eggs can be fertilized through natural matings or by in vitro fertilization and examined visually. Here we review how eggs are obtained and how to recognize healthy eggs that will develop into high-quality embryos.

Adult frogs that are well-cared-for will give high-quality eggs and embryos for use in everyXenopusprotocol. Thoughtful frog husbandry is thus pivotal to successful research using these organisms. Protocols for successfully raising tadpoles, establishing and maintaining water quality, and detecting specific pathogens are key to maintaining healthy frog populations.

In people and other vertebrates, craniofacial development leads to formation of the skull, facial bones, and cartilage. Craniofacial development is complex, involving multiple cell types, with carefully coordinated cell movements, and is often disrupted, with disorders occurring in ~1/700 of live human births. The frog Xenopus has proven an outstanding model for understanding mechanisms of craniofacial development. Neural crest cells are unique to vertebrates and contribute a large part of the craniofacial skeleton. The mouth is conserved and essential, arising from ectodermal and endodermal cells. Multiple signaling pathways and regulatory genes are required for craniofacial development, including a major signaling center, the extreme anterior domain. Present treatments of craniofacial disorders are largely surgical. The need for definition of genes and molecular mechanisms underlying craniofacial disorders is critical and an area in which Xenopus is making key contributions. Xenopus remains one of the most accessible systems for uncovering connections to human craniofacial disorders.

The complex 16p11.2 Deletion Syndrome (16pdel) is accompanied by neurological disorders, including epilepsy, autism spectrum disorder and intellectual disability. We demonstrate that 16pdel iPSC differentiated neurons showed augmented local field potential activity and altered ceramide-related lipid species relative to unaffected. FAM57B , a poorly characterized gene in the 16p11.2 interval, has emerged as a candidate tied to symptomatology. We found that FAM57B modulates ceramide synthase (CerS) activity, but is not a CerS per se. In FAM57B mutant human neuronal cells and zebrafish brain, composition and levels of sphingolipids and glycerolipids associated with cellular membranes are disrupted. Consistently, we observed aberrant plasma membrane architecture and synaptic protein mislocalization, which were accompanied by depressed brain and behavioral activity. Together, these results suggest that haploinsufficiency of FAM57B contributes to changes in neuronal activity and function in 16pdel Syndrome, through a crucial role for the gene in lipid metabolism.

Zebrafish are an important and expanding experimental system for brain research. We describe a noninvasive electrophysiology technique that can be used in living larvae to measure spontaneous activity in the brain and spinal cord simultaneously. This easy-to-use method uses a commercially available multielectrode array to detect local field potential parameters, and allows for relative coordinated (network) measurements of activity. We demonstrate sensitivity of this system by measuring activity in larvae treated with the antiepileptic drug valproic acid. Valproic acid decreased larval movement and startle response, and decreased spontaneous brain activity. Spinal cord activity did not change after treatment, suggesting valproic acid primarily affects brain function. The observed differences in brain activity, but not spinal cord activity, after valproic acid treatment indicates that brain activity differences are not a secondary effect of decreased startle response and movement. We provide a step-by-step protocol for experiments presented that a novice could easily follow. This electrophysiological method will be useful to the zebrafish neuroscience community.

The brain ventricular system is a series of connected cavities, filled with cerebrospinal fluid (CSF), that forms within the vertebrate central nervous system (CNS). The hollow neural tube is a hallmark of the chordate CNS, and a closed neural tube is essential for normal development. Development and function of the ventricular system is examined, emphasizing three interdigitating components that form a functional system: ventricle walls, CSF fluid properties, and activity of CSF constituent factors. The cellular lining of the ventricle both can produce and is responsive to CSF. Fluid properties and conserved CSF components contribute to normal CNS development. Anomalies of the CSF/ventricular system serve as diagnostics and may cause CNS disorders, further highlighting their importance. This review focuses on the evolution and development of the brain ventricular system, associated function, and connected pathologies. It is geared as an introduction for scholars with little background in the field.

Basal constriction occurs at the zebrafish midbrain-hindbrain boundary constriction (MHBC) and is likely a widespread morphogenetic mechanism. 3D reconstruction demonstrates that MHBC cells are wedge-shaped, and initially constrict basally, with subsequent apical expansion. wnt5b is expressed in the MHB and is required for basal constriction. Consistent with a requirement for this pathway, expression of dominant negative Gsk3β overcomes wnt5b knockdown. Immunostaining identifies focal adhesion kinase (Fak) as active in the MHB region, and knockdown demonstrates Fak is a regulator of basal constriction. Tissue specific knockdown further indicates that Fak functions cell autonomously within the MHBC. Fak is epistatic to wnt5b , suggesting that Wnt5b signals locally as an early step in basal constriction and acts together with more widespread Fak activation. This study delineates signaling pathways that regulate basal constriction during brain morphogenesis.

Basal constriction occurs at the zebrafish midbrain–hindbrain boundary constriction (MHBC) and is likely a widespread morphogenetic mechanism. 3D reconstruction demonstrates that MHBC cells are wedge-shaped, and initially constrict basally, with subsequent apical expansion. wnt5b is expressed in the MHB and is required for basal constriction. Consistent with a requirement for this pathway, expression of dominant negative Gsk3β overcomes wnt5b knockdown. Immunostaining identifies focal adhesion kinase (Fak) as active in the MHB region, and knockdown demonstrates Fak is a regulator of basal constriction. Tissue specific knockdown further indicates that Fak functions cell autonomously within the MHBC. Fak acts downstream of wnt5b, suggesting that Wnt5b signals locally as an early step in basal constriction and acts together with more widespread Fak activation. This study delineates signaling pathways that regulate basal constriction during brain morphogenesis.