Biology subjects

Fletcher, J. L.

Publications and source records attributed to Fletcher, J. L..

5 recordsLinked to original sources

bioRxiv2024

Gluk4-containing kainate receptors regulate synaptic communication in the motor cortex and reduce axon degeneration in adult mice

Glutamate-gated kainate receptors comprising the Gluk4 subunit (encoded by Grik4) are highly expressed by neurons in the central nervous system. We report that Grik4 mRNA is widely expressed by neurons in the adult mouse motor cortex, where GluK4-containing kainate receptors account for [~]60% of the kainate evoked current in layer V pyramidal neurons. To elucidate their role in motor circuit regulation, we analysed the behaviour of mice that lacked the pore forming domain of the GluK4 subunit (Grik4-/-mice). Grik4-/- mice were hyperactive, had an abnormal gait, and impaired motor coordination. At postnatal day (P)60, layer V pyramidal neurons received fewer miniature excitatory post synaptic currents, had a reduced density of thin spines on their basal dendrites, and a reduced density of VGlut1 puncta at the soma, consistent with neurons receiving fewer excitatory synaptic connections. Grik4-/- mice also lost [~]44% of their callosal axons between P60 and P180 and the amplitude of the callosal compound action potential was reduced by [~]25-30%. RNA sequencing data support the capacity for Grik4 to modulate synaptic and neuroprotective signalling pathways.

neuroscience↗

bioRxiv2023

Induced pluripotent stem cell derived pericytes respond to endogenous mediators of proliferation and contractility

BackgroundPericytes are multifunctional contractile cells that reside on capillaries. Pericytes are critical regulators of cerebral blood flow and blood-brain barrier function, and pericyte dysfunction may contribute to the pathophysiology of human neurological diseases including Alzheimers disease, multiple sclerosis, and stroke. Induced pluripotent stem cell (iPSC)-derived pericytes (iPericytes) are a promising tool for vascular research. However, it is unclear how iPericytes functionally compare to primary human brain vascular pericytes (HBVPs). We differentiated iPSCs into iPericytes of either the mesoderm or neural crest lineage using established protocols. We compared iPericyte and HBVP morphologies, quantified gene expression by qPCR and bulk RNA sequencing, and visualised pericyte protein markers by immunocytochemistry. To determine whether the gene expression of neural crest iPericytes, mesoderm iPericytes or HBVPs correlated with their functional characteristics in vitro, we quantified EdU incorporation following exposure to the key pericyte mitogen, platelet derived growth factor (PDGF)-BB and, contraction and relaxation in response to the vasoconstrictor endothelin-1 or vasodilator adenosine, respectively. iPericytes were morphologically similar to HBVPs and expressed canonical pericyte markers. However, iPericytes had 1864 differentially expressed genes compared to HBVPs, while there were 797 genes differentially expressed between neural crest and mesoderm iPericytes. Consistent with the ability of HBVPs to respond to PDGF-BB signalling, PDGF-BB enhanced and PDGF receptor-beta inhibitors impaired iPericyte proliferation. Administration of endothelin-1 led to iPericyte contraction and adenosine led to iPericyte relaxation, of a magnitude similar to the response evoked in HBVPs. We determined that neural crest iPericytes were less susceptible to PDGFR beta inhibition, but responded most robustly to vasoconstrictive meditators. iPericytes express pericyte-associated genes and proteins and, exhibit an appropriate physiological response upon exposure to a key endogenous mitogen or vasoactive mediators. Therefore, the generation of functional iPericytes would be suitable for use in future investigations exploring pericyte function or dysfunction in neurological diseases.

cell biology↗

bioRxiv2021

Central nervous system infusion of TrkB agonist, 7,8-dihydroxyflavone, is ineffective in promoting myelin repair in cuprizone and experimental autoimmune encephalomyelitis mouse models of multiple sclerosis

Small molecular weight functional mimetics of brain-derived neurotrophic factor (BDNF) which act via the TrkB receptor have been developed to overcome the pharmacokinetic limitations of BDNF as a therapeutic for neurological disease. Activation of TrkB on oligodendrocytes has been identified as a potential strategy for myelin repair in demyelinating conditions. Here, we tested the efficacy of intracerebroventricular infusion of TrkB agonist 7,8-dihydroxyflavone (DHF) to promote myelin repair in the cuprizone model and alter the course of experimental autoimmune encephalomyelitis (EAE). In these two distinct mouse models used for the preclinical testing of remyelinating therapeutics, we found that DHF infusion increased the percentage of myelin basic protein and density of oligodendrocyte progenitor cells (OPCs) in the corpus callosum of female C57BL/6 mice after cuprizone demyelination. However, DHF did not alter the percentage of axons myelinated or increase the density of post-mitotic oligodendrocytes in this model. Direct cerebrospinal fluid infusion of DHF also had no effect on the clinical course of EAE, and examination of the lumbar spinal cord after 21 days of treatment revealed extensive demyelination. These results indicate that direct cerebrospinal fluid infusion of DHF is ineffective at promoting myelin repair in toxin-induced and inflammatory models of demyelination.

neuroscience↗

bioRxiv2020

Acute treatment with TrkB agonist LM22A-4 confers neuroprotection and preserves myelin integrity in a mouse model of pediatric traumatic brain injury

Young children have a high risk of sustaining a traumatic brain injury (TBI), which can have debilitating life-long consequences. Importantly, the young brain shows particular vulnerability to injury, likely attributed to ongoing maturation of the myelinating nervous system at the time of insult. Here, we examined the effect of acute treatment with partial tropomyosin receptor kinase B (TrkB) agonist, LM22A-4, on the pathological and neurobehavioral outcomes after pediatric TBI, with the hypothesis that targeting TrkB would minimize tissue damage and support functional recovery. We focused on myelinated tracts-- the corpus callosum and external capsules--based on recent evidence that TrkB activation potentiates oligodendrocyte remyelination. Male mice at postnatal day 21 received an experimental TBI or sham surgery. Acutely post-injury, extensive cell death, a robust glial response and disruption of compact myelin were evident in the injured brain. TBI or sham mice then received intranasal saline vehicle or LM22A-4 for 14 days. Behavior testing was performed from 4 weeks post-injury, and brains were collected at 5 weeks for histology. TBI mice showed hyperactivity, reduced anxiety-like behavior, and social memory impairments. LM22A-4 ameliorated the abnormal anxiolytic phenotype but had no effect on social memory deficits. Use of spectral confocal reflectance microscopy detected persistent myelin fragmentation in the external capsule of TBI mice at 5 weeks post-injury, which was accompanied by regionally distinct deficits in oligodendrocyte progenitor cells and postmitotic oligodendrocytes, as well as chronic reactive gliosis and atrophy of the corpus callosum and injured external capsule. LM22A-4 treatment ameliorated myelin deficits in the perilesional external capsule, as well as tissue volume loss and the extent of reactive gliosis. However, there was no effect of this TrkB agonist on oligodendroglial populations detected at 5 weeks post-injury. Collectively, our results demonstrate that targeting TrkB immediately after TBI during early life confers neuroprotection and preserves myelin integrity, and this was associated with some improved neurobehavioral outcomes as the pediatric injured brain matures.

neuroscience↗

bioRxiv2018

Targeting TrkB with a brain-derived neurotrophic factor mimetic promotes myelin repair in the brain

Methods to promote myelin regeneration in response to central myelin loss are essential to prevent the progression of clinical disability in demyelinating diseases. The neurotrophin brain-derived neurotrophic factor (BDNF) is known to promote myelination during development via oligodendrocyte expressed TrkB receptors. Here, we use a structural mimetic of BDNF to promote myelin regeneration in a preclinical mouse model of central demyelination. We show that selective targeting of TrkB with the BDNF-mimetic enhances remyelination, increasing oligodendrocyte differentiation, the frequency of myelinated axons, and myelin sheath thickness after a demyelinating insult. Treatment with exogenous BDNF exerted an attenuated effect, increasing myelin sheath thickness only. Further, following conditional deletion of TrkB from pre-myelinating oligodendrocytes, we show the effects of the BDNF-mimetic on oligodendrocyte differentiation and remyelination are lost, indicating these are dependent on oligodendrocyte expression of TrkB. Overall, these studies demonstrate that targeting oligodendrocyte TrkB promotes in vivo remyelination in the brain.

neuroscience↗