Calcium Imaging of Neuronal Activity in Drosophila Can Identify Anticonvulsive Compounds

PLoS One. 2016 Feb 10;11(2):e0148461. doi: 10.1371/journal.pone.0148461. eCollection 2016.

Abstract

Although there are now a number of antiepileptic drugs (AEDs) available, approximately one-third of epilepsy patients respond poorly to drug intervention. The reasons for this are complex, but are probably reflective of the increasing number of identified mutations that predispose individuals to this disease. Thus, there is a clear requirement for the development of novel treatments to address this unmet clinical need. The existence of gene mutations that mimic a seizure-like behaviour in the fruit fly, Drosophila melanogaster, offers the possibility to exploit the powerful genetics of this insect to identify novel cellular targets to facilitate design of more effective AEDs. In this study we use neuronal expression of GCaMP, a potent calcium reporter, to image neuronal activity using a non-invasive and rapid method. Expression in motoneurons in the isolated CNS of third instar larvae shows waves of calcium-activity that pass between segments of the ventral nerve cord. Time between calcium peaks, in the same neurons, between adjacent segments usually show a temporal separation of greater than 200 ms. Exposure to proconvulsants (picrotoxin or 4-aminopyridine) reduces separation to below 200 ms showing increased synchrony of activity across adjacent segments. Increased synchrony, characteristic of epilepsy, is similarly observed in genetic seizure mutants: bangsenseless1 (bss1) and paralyticK1270T (paraK1270T). Exposure of bss1 to clinically-used antiepileptic drugs (phenytoin or gabapentin) significantly reduces synchrony. In this study we use the measure of synchronicity to evaluate the effectiveness of known and novel anticonvulsive compounds (antipain, isethionate, etopiside rapamycin and dipyramidole) to reduce seizure-like CNS activity. We further show that such compounds also reduce the Drosophila voltage-gated persistent Na+ current (INaP) in an identified motoneuron (aCC). Our combined assays provide a rapid and reliable method to screen unknown compounds for potential to function as anticonvulsants.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 4-Aminopyridine / pharmacology
  • Adult
  • Amines / pharmacology
  • Animals
  • Anticonvulsants / chemical synthesis
  • Anticonvulsants / pharmacology*
  • Antipain / pharmacology
  • Calcium / metabolism*
  • Calmodulin / genetics
  • Calmodulin / metabolism
  • Central Nervous System / cytology
  • Central Nervous System / drug effects
  • Central Nervous System / metabolism
  • Convulsants / pharmacology
  • Cyclohexanecarboxylic Acids / pharmacology
  • Dipyridamole / pharmacology
  • Drosophila Proteins / genetics
  • Drosophila Proteins / metabolism
  • Drosophila melanogaster / cytology
  • Drosophila melanogaster / drug effects*
  • Drosophila melanogaster / metabolism
  • Female
  • Gabapentin
  • Gene Expression
  • Genes, Reporter
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • High-Throughput Screening Assays*
  • Humans
  • Larva / cytology
  • Larva / drug effects
  • Larva / metabolism
  • Male
  • Molecular Imaging / methods
  • Motor Neurons / cytology
  • Motor Neurons / drug effects*
  • Motor Neurons / metabolism
  • Phenytoin / pharmacology
  • Picrotoxin / pharmacology
  • Primary Cell Culture
  • Sodium Channels / genetics
  • Sodium Channels / metabolism
  • gamma-Aminobutyric Acid / pharmacology

Substances

  • Amines
  • Anticonvulsants
  • Calmodulin
  • Convulsants
  • Cyclohexanecarboxylic Acids
  • Drosophila Proteins
  • Sodium Channels
  • para protein, Drosophila
  • Picrotoxin
  • Green Fluorescent Proteins
  • Antipain
  • gamma-Aminobutyric Acid
  • Phenytoin
  • Dipyridamole
  • Gabapentin
  • 4-Aminopyridine
  • Calcium