Biology subjects

Kanyo, R.

Publications and source records attributed to Kanyo, R..

4 recordsLinked to original sources

bioRxiv2026

Pharmacological profiling of brain activity in zebrafish

Background and PurposeEpilepsy is a neurological condition characterized by recurring seizures and neuronal hyperexcitability. Cell-based high-throughput screening applications have been essential for drug development and discovering novel biological processes. However, cell-based screens do not provide information on how drug-targeted pathways are integrated into a whole animal. Our objective was to develop and evaluate a screening application using zebrafish larvae to identify signalling mechanisms that modulate neural activity. Experimental ApproachWe developed an in vivo automated high-content screening assay using zebrafish larvae expressing the calcium sensor CaMPARI (calcium-modulated photoactivatable ratiometric integrator) in neurons. This assay can quantify neural activity of multiple individual larvae per well in a 96-well format. We quantified neural activity in 8725 individual larvae, in response to 1292 different drugs to identify molecules that protect against convulsant-induced neuronal hyperexcitability. Key ResultsThe assay was effective at identifying drugs that target diverse neurotransmitter signalling systems. While some commonly used anti-convulsants (e.g. phenytoin, carbamazepine, valproic acid) had poor activity in the assay, Kv7 potassium channel activators were consistently effective (ICA-069673, ICA-27243, ICA-110381, retigabine, and ML213). Many compounds approved for treatment of other conditions, including amitriptyline (depression), cyclobenzaprine (muscle spasm), clomipramine (obsessive-compulsive disorder) and ganaxolone (seizures), also strongly suppressed excitability in the assay. Conclusion and ImplicationsNeuronal CaMPARI expression in zebrafish larvae is a powerful tool for plate-based compound library screening to identify drugs that suppress hyperexcitability in vivo. Bullet Point SummaryO_ST_ABSWhat is already knownC_ST_ABSO_LICaMPARI is an integrative Ca2+ sensor that can be used to identify active neurons. C_LIO_LIKv7 activators (retigabine, ML213, and ICA-069673) are effective at reducing convulsant-induced (4-AP) neuronal hyperexcitability. C_LI What this study addsO_LIAn automated in vivo high-content drug screening assay to quantify neural activity. C_LIO_LIA series of drug targets that influence convulsant-induced hyperexcitability. C_LI Clinical significanceO_LIOur new tool will help identify novel compounds and signalling mechanisms that could be pursued as therapeutic targets for diseases involving electrical hyperexcitability. C_LI

pharmacology and toxicology↗

bioRxiv2021

Embryonic exposure to cannabidiol disrupts active neural circuits, an effect increased by Δ-tetrahydrocannabinol and involving CB1R and CB2R in zebrafish

In the light of legislative changes and the widespread use of cannabis as a recreational and medicinal drug, delayed effects of cannabis upon brief exposure during embryonic development are of high interest as early pregnancies often go undetected. Here, zebrafish embryos were exposed to cannabidiol (CBD) and {Delta}-9-tetrahydrocannabinol l (THC) until the end of gastrulation (1-10 hours post-fertilization) and analyzed later in development (4-5 days post-fertilization). In order to measure neural activity, we implemented CaMPARI (Calcium-Modulated Photoactivatable Ratiometric Integrator) and optimized the protocol for a 96-well format complemented by locomotor analysis. Our results revealed that neural activity was decreased by CBD more than THC. At higher doses, both cannabinoids could dramatically reduce neural activity and locomotor activity. Interestingly, the decrease was more pronounced when CBD and THC were combined. At the receptor level, CBD-mediated reduction of locomotor activity was partially prevented using cannabinoid type 1 and 2 receptor inhibitors. Overall, we report that CBD toxicity occurs via two cannabinoid receptors and is synergistically enhanced by THC exposure to negatively impact neural activity late in larval development. Future studies are warranted to reveal other cannabinoids and receptors involved in this pathway to understand the subsequent health implications of cannabis consumption on fetal development.

pharmacology and toxicology↗

bioRxiv2020

Seizures are a druggable mechanistic link between TBI and subsequent tauopathy

Traumatic brain injury (TBI) is a prominent risk factor for neurodegenerative diseases and dementias including chronic traumatic encephalopathy (CTE). TBI and CTE, like all tauopathies, are characterized by accumulation of Tau into aggregates that progressively spread to other brain regions in a prion-like manner. The mechanisms that promote spreading and cellular uptake of tau seeds after TBI are not fully understood, in part due to lack of tractable animal models. Here, we test the putative roles for excess neuronal activity and dynamin-dependent endocytosis in promoting the in vivo spread of tauopathy. We introduce tauopathy reporter zebrafish expressing a genetically-encoded fluorescent Tau biosensor that reliably reports accumulation of human tau species when seeded via intra-ventricular brain injections. Subjecting zebrafish larvae to a novel TBI paradigm produced various TBI symptoms including cell death, hemorrhage, blood flow abnormalities, post-traumatic seizures, and Tau inclusions. Bath application of anticonvulsant drugs rescued TBI-induced tauopathy and cell death; these benefits were attributable to inhibition of post-traumatic seizures because co-application of convulsants reversed these beneficial effects. However, one convulsant drug, 4-Aminopyridine, unexpectedly abrogated TBI-induced tauopathy - this was due to its inhibitory action on endocytosis as confirmed via additional dynamin inhibitors. These data suggest a role for seizure activity and dynamin-dependent endocytosis in the prion-like seeding and spreading of tauopathy following TBI. Further work is warranted regarding anti-convulsants that dampen post-traumatic seizures as a route to moderating subsequent tauopathy. Moreover, the data highlight the utility of deploying in vivo Tau biosensor and TBI methods in larval zebrafish, especially regarding drug screening and intervention. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=117 SRC="FIGDIR/small/091819v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@34a2caorg.highwire.dtl.DTLVardef@1ab916borg.highwire.dtl.DTLVardef@d83bf2org.highwire.dtl.DTLVardef@47bf90_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIIntroduces first Traumatic Brain Injury (TBI) model in larval zebrafish, and its easy C_LIO_LITBI induces clinically relevant cell death, haemorrhage & post-traumatic seizures C_LIO_LICa2+ imaging during TBI reveals spike in brain activity concomitant with seizures C_LIO_LITau-GFP Biosensor allows repeated in vivo measures of prion-like tau aggregation C_LIO_LIpost-TBI, anticonvulsants stop tauopathies akin to Chronic Traumatic Encephalopathy C_LI

neuroscience↗

bioRxiv2020

A bright and high-performance genetically encoded Ca2+ indicator based on mNeonGreen fluorescent protein

Genetically encodable calcium ion (Ca2+) indicators (GECIs) based on green fluorescent proteins (GFP) are powerful tools for imaging of cell signaling and neural activity in model organisms. Following almost two decades of steady improvements in the Aequorea victoria GFP (avGFP)-based GCaMP series of GECIs, the performance of the most recent generation (i.e., GCaMP7) may have reached its practical limit due to the inherent properties of GFP. In an effort to sustain the steady progression towards ever-improved GECIs, we undertook the development of a new GECI based on the bright monomeric GFP, mNeonGreen (mNG). The resulting indicator, mNG-GECO1, is 60% brighter than GCaMP6s in vitro and provides comparable performance as demonstrated by imaging Ca2+ dynamics in cultured cells, primary neurons, and in vivo in larval zebrafish. These results suggest that mNG-GECO1 is a promising next-generation GECI that could inherit the mantle of GCaMP and allow the steady improvement of GECIs to continue for generations to come.

molecular biology↗