Transcriptomes of post-mitotic neurons identify the usage of alternative pathways during adult and embryonic neuronal differentiation

Original Panther analysis tool data tables showing values and p -values for overrepresentation data sets. (XLSX 21 kb

The Met receptor tyrosine kinase prevents zebrafish primary motoneurons from expressing an incorrect neurotransmitter

Abstract Background Expression of correct neurotransmitters is crucial for normal nervous system function. How neurotransmitter expression is regulated is not well-understood; however, previous studies provide evidence that both environmental signals and intrinsic differentiation programs are involved. One environmental signal known to regulate neurotransmitter expression in vertebrate motoneurons is Hepatocyte growth factor, which acts through the Met receptor tyrosine kinase and also affects other aspects of motoneuron differentiation, including axonal extension. Here we test the role of Met in development of motoneurons in embryonic zebrafish. Results We found that met is expressed in all early developing, individually identified primary motoneurons and in at least some later developing secondary motoneurons. We used morpholino antisense oligonucleotides to knock down Met function and found that Met has distinct roles in primary and secondary motoneurons. Most secondary motoneurons were absent from met morpholino-injected embryos, suggesting that Met is required for their formation. We used chemical inhibitors to test several downstream pathways activated by Met and found that secondary motoneuron development may depend on the p38 and/or Akt pathways. In contrast, primary motoneurons were present in met morpholino-injected embryos. However, a significant fraction of them had truncated axons. Surprisingly, some CaPs in met morpholino antisense oligonucleotide (MO)-injected embryos developed a hybrid morphology in which they had both a peripheral axon innervating muscle and an interneuron-like axon within the spinal cord. In addition, in met MO-injected embryos primary motoneurons co-expressed mRNA encoding Choline acetyltransferase, the synthetic enzyme for their normal neurotransmitter, acetylcholine, and mRNA encoding Glutamate decarboxylase 1, the synthetic enzyme for GABA, a neurotransmitter never normally found in these motoneurons, but found in several types of interneurons. Our inhibitor studies suggest that Met function in primary motoneurons may be mediated through the MEK1/2 pathway. Conclusion We provide evidence that Met is necessary for normal development of zebrafish primary and secondary motoneurons. Despite their many similarities, our results show that these two motoneuron subtypes have different requirements for Met function during development, and raise the possibility that Met may act through different intracellular signaling cascades in primary and secondary motoneurons. Surprisingly, although met is not expressed in primary motoneurons until many hours after they have extended axons to and innervated their muscle targets, Met knockdown causes some of these cells to develop a hybrid phenotype in which they co-expressed motoneuron and interneuron neurotransmitters and have both peripheral and central axons.</p

Pilot Screening to Identify Social Circuitry

Project files include 1 page pdf.Humans have a long history of working in social groups with a plethora of research supporting the benefits of positive human interaction. While these behaviors may seem intuitive, they can be argued as the most intricate behaviors displayed by animals. To generate a situationally relevant response to social others, an animal must be able to track dynamic systems and recall past experiences. Given the inherent multisensory nature of social behaviors, it is expected that this system can be perturbed by a multitude of neurological disorders. Using animal models, we can begin to construct the neuronal circuitry necessary for development of social behaviors. This circuitry can be used to understand social behavior deficits and may reveal possible interventions for disorders. Zebrafish, Danio rerio, perform stereotyped social behaviors, such as shoaling, which can be used to explore neuronal changes associated with deficits in performing the behavior. Through chemo-genetic ablations, we were able to cause cell death to select neurons due to variations in gene expression. Once chemically treated, fish were put into a virtual social assay that untreated zebrafish respond to. Zebrafish exhibiting decreased social response would indicate that the ablated neurons are necessary for social behavior

Additional file 1 of A conserved transcriptional fingerprint of multi-neurotransmitter neurons necessary for social behavior

Additional file 1: Table 1. LIMTF Gene expression in vTely321 neurons. Key Resource Table 1. Key Resource Table 2. Additional Fig. 1.chatb labelling in vTely321 neurons. Validation of cholinergic identity of vTely321 neurons with chatb ISH probes. White arrows point to selected vTely321 neurons expressing both chatb and GABA. (Scale bars = 20 μm). Additional Fig. 2.gad1b&2 labelling validates GABA immunostaining. gad1b and gad2 transcripts were labelled in vTely321 neurons and validation staining was performed with GABA antibody. White arrows denote GABA and gad1b&2 double-labeling, Yellow arrows label vTely321 neurons which are positive for both GABA and gad1b&2 in the main figure and insert. GABA antibody labelling overlaps extensively with gad1b&2 confirming its specificity. (Scale bars = 20 μm). Additional Fig. 3. GABA antibody labels GABAergic neurons in the larval spinal cord. To test the specificity of the GABA antibody we imaged 24–48 hpf larval spinal cords of the transgenic line Tg (mnx1:GAL4; UAS:GFP) which drives GFP expression (yellow) in cholinergic motor neurons. White arrow denotes KA neurons and Cyan arrow points to DoLA neurons which are known GABAergic neurons in the spinal cord. Yellow arrows denote motor neurons (MN). The absence of overlap between the MN and cyan labelling confirms specificity of the GABA antibody. (Scale bars represent 20 μm)

Harmonin (Ush1c) is required in zebrafish Muller glial cells for photoreceptor synaptic development and function

Usher syndrome is the most prevalent cause of hereditary deaf-blindness, characterized by congenital sensorineural hearing impairment and progressive photoreceptor degeneration beginning in childhood or adolescence. Diagnosis and management of this disease are complex, and the molecular changes underlying sensory cell impairment remain poorly understood. Here we characterize two zebrafish models for a severe form of Usher syndrome, Usher syndrome type 1C (USH1C): one model is a mutant with a newly identified ush1c nonsense mutation, and the other is a morpholino knockdown of ush1c. Both have defects in hearing, balance and visual function from the first week of life. Histological analyses reveal specific defects in sensory cell structure that are consistent with these behavioral phenotypes and could implicate Müller glia in the retinal pathology of Usher syndrome. This study shows that visual defects associated with loss of ush1c function in zebrafish can be detected from the onset of vision, and thus could be applicable to early diagnosis for USH1C patients

Harmonin (Ush1c) is required in zebrafish Müller glial cells for photoreceptor synaptic development and function

SUMMARY Usher syndrome is the most prevalent cause of hereditary deaf-blindness, characterized by congenital sensorineural hearing impairment and progressive photoreceptor degeneration beginning in childhood or adolescence. Diagnosis and management of this disease are complex, and the molecular changes underlying sensory cell impairment remain poorly understood. Here we characterize two zebrafish models for a severe form of Usher syndrome, Usher syndrome type 1C (USH1C): one model is a mutant with a newly identified ush1c nonsense mutation, and the other is a morpholino knockdown of ush1c. Both have defects in hearing, balance and visual function from the first week of life. Histological analyses reveal specific defects in sensory cell structure that are consistent with these behavioral phenotypes and could implicate Müller glia in the retinal pathology of Usher syndrome. This study shows that visual defects associated with loss of ush1c function in zebrafish can be detected from the onset of vision, and thus could be applicable to early diagnosis for USH1C patients