title: A comparison of optical and electrophysiological methods for recording retinal ganglion cells during electrical stimulation.
abstract: OBJECTIVE
  To compare the efficacy of optical techniques with electrophysiological recordings for mapping retinal activity in response to electrical stimulation.
  
  
  METHODS
  Whole cell patch clamp, Ca(2+) imaging (Fluo-4-AM), and Na(+) imaging (CoroNa Green-AM) techniques were used to detect responses of neurons from mouse and salamander retina to electrical stimulation.
  
  
  RESULTS
  Synaptic currents were observed in ≥23% of retinal ganglion cells (RGCs), indicating presynaptic Ca(2+) increases in the inner plexiform layer (IPL). Modest depolarization with 20-30 mM K(+) consistently evoked Ca(2+) responses measured with Fluo4, but Ca(2+) responses were almost never evoked by epiretinal stimulation. In salamander retina, responses were seen in the inner nuclear layer (INL) and IPL. In mouse retina, responses were also sometimes seen in the outer pexiform layer (OPL). OPL responses showed a longer latency than IPL responses, suggesting that outer retinal circuits do not trigger synaptic responses of RGCs. Simultaneous Ca(2+) imaging and electrophysiological recording of synaptic currents confirmed that Fluo4-loaded retinas remained responsive to stimulation. Epiretinal stimulation evoked action potentials in ≥67% of RGCs. CoroNa Green detected Na(+) changes stimulated by 20 mM K(+), but epiretinal stimulation did not evoke detectable Na(+) responses. Simultaneous imaging and electrophysiological recording confirmed the health of CoroNa Green-loaded retinas. We confirmed stimulation efficacy by simultaneously recording Na(+) changes and electrophysiological responses.
  
  
  CONCLUSIONS
  These data demonstrate that electrophysiological recordings show greater sensitivity than Na(+) or Ca(2+) imaging in response to electrical stimulation. The paucity of Ca(2+) responses is consistent with limited risk for Ca(2+)-mediated cell damage during electrical stimulation.
authors.affiliation: Department of Ophthalmology and Visual Science, University of Nebraska Medical Center, Omaha, NE 68198, USA.
authors.forename: Jianmin
authors.initials: J
authors.lastname: Luo
authors.forename: Bronson J
authors.initials: BJ
authors.lastname: Boosalis
authors.forename: Wallace B
authors.initials: WB
authors.lastname: Thoreson
authors.forename: Eyal
authors.initials: E
authors.lastname: Margalit
mesh_terms: D000200:Action Potentials / Q000502:physiology*; D000557:Ambystoma; D000814:Aniline Compounds / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D004558:Electric Stimulation; D004594:Electrophysiology / Q000379:methods; D005456:Fluorescent Dyes / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018613:Microscopy, Confocal / Q000379:methods; D018408:Patch-Clamp Techniques; D012165:Retinal Ganglion Cells / Q000502:physiology*; D012964:Sodium / Q000378:metabolism; D055366:Synaptic Potentials / Q000502:physiology*; D014966:Xanthenes / Q000378:metabolism
chemical_list: D000814:Aniline Compounds; C409648:Fluo 4; D005456:Fluorescent Dyes; D014966:Xanthenes; D012964:Sodium; D002118:Calcium