Voltage-gated sodium channels (Na v channels) in retinal neurons are known to contribute to the mammalian flash electroretinogram (ERG) via activity of third-order retinal neurons, i.e. amacrine and ganglion cells. This study investigated the effects of tetrodotoxin (TTX) blockade of Na v channels on the b-wave, an ERG wave that originates mainly from activity of second-order retinal neurons. ERGs were recorded from anaesthetized Brown Norway rats in response to brief full-field flashes presented over a range of stimulus energies, under dark-adapted conditions and in the presence of steady mesopic and photopic backgrounds. Recordings were made before and after intravitreal injection of TTX (∼3 μM) alone, 3-6 weeks after optic nerve transection (ONTx) to induce ganglion cell degeneration, or in combination with an ionotropic glutamate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 200 μM) to block light-evoked activity of inner retinal, horizontal and OFF bipolar cells, or with the glutamate agonist N -methyl-D-aspartate (NMDA, 100-200 μM) to reduce light-evoked inner retinal activity. TTX reduced ERG amplitudes measured at fixed times corresponding to b-wave time to peak. Effects of TTX were seen under all background conditions, but were greatest for mesopic backgrounds. In dark-adapted retina, b-wave amplitudes were reduced only when very low stimulus energies affecting the inner retina, or very high stimulus energies were used. Neurophysiological studies of blockade of Na v channels using tetrodotoxin (TTX) (Narahashi et al. 1964;Kao, 1966) in isolated mammalian retinal slice preparations or dissociated neurons have shown that these channels are prominent and functional in retinal ganglion cells (e.g. Lipton & Tauck, 1987;Skaliora et al. 1993) and in some This paper has online supplemental material. Loss of ganglion cells following ONTx did not affect b-wave amplitudes, and injection of TTX in eyes withtypes of amacrine cells (e.g. AII, A17, A18 and starburst amacrine cells; Boos et al. 1993;Feigenspan et al. 1998;Hartveit, 1999;Cohen, 2001). More recently, functional Na v channels in isolated bipolar cells and bipolar cells in retinal slice preparations have been observed in several species: rat (Pan & Hu, 2000), goldfish (Zenisek et al. 2001), salamander Ichinose & Lukasiewicz, 2007), ground squirrel (Saszik & DeVries, 2005) human (Miyachi et al. 2006;Ohkuma et al. 2007). In the rat retina, functional Na v channels were found in cone but not rod bipolar cells (Pan & Hu, 2000). Their absence from rod bipolar cells, suggests an additional postreceptoral divergence in physiological properties of rod versus cone circuits (Pan & Hu, 2000;Ma et al. 2005). The activity of Na v channels in bipolar cells has been shown recently to be modulated by ambient background illumination in the tiger salamander retinal slice (Ichinose & Lukasiewicz, 2007) where the Na v channels contributed more to the bipolar cell response under low light conditions than high light conditions. The authors suggested that the ...

RGS7, RGS11, and their binding partner G␤5 are localized to the dendritic tips of retinal ON bipolar cells (ON-BPC), where mGluR6 responds to glutamate released from photoreceptor terminals by activation of the RGS7/RGS11 substrate, G␣o. To determine their functions in retinal signaling, we investigated cell-specific expression patterns of RGS7 and RGS11 by immunostaining, and measured light responses by electroretinography in mice with targeted disruptions of the genes encoding them. RGS7 staining is present in dendritic tips of all rod ON-BPC, but missing in those for subsets of cone ON-BPC, whereas the converse was true for RGS11 staining. Genetic disruption of either RGS7 or RGS11 produced delays in the ON-BPC-derived electroretinogram b-wave, but no changes in the photoreceptor-derived a-wave. Homozygous RGS7 mutant mice had delays in rod-driven b-waves, whereas RGS11 mutant mice had delays in rod-driven, and especially in cone-driven b-waves. The b-wave delays were further enhanced in mice homozygous for both RGS7 and RGS11 gene disruptions. Thus, RGS7 and RGS11 act in parallel to regulate the kinetics of ON bipolar cell responses, with differential impacts on the rod and cone pathways.

Introduction:Measurement of static pupillary size in the ICU is of importance in cases of acutely expanding intracranial mass lesions. The inaccuracies with subjective assessment of pupillary size by medical personnel preclude its use in emergent neurological situations.Objective:To determine if the ratio of pupil to limbus diameter (PLD ratio) measured by a two-box method is a reliable measure of pupil size for detecting early anisocoria and measuring pupillary changes.Materials and Methods:The PLD ratio was defined as the ratio of the pupillary diameter measured at a para-horizontal axial plane with the limbus diameter measured at the same or parallel axial plane. A two-box method was used to estimate the diameters of imaged pupils. Eyes were imaged using an iPhone 4S cellphone camera. Background illumination was measured and kept constant. The pupils of a 78-year-old woman, who presented with a large intra-axial parenchymal hemorrhage, were imaged. The patient had left pupillary miosis in dark but not in bright light. After presenting this case along with the images of the pupillary examination, a group of 21 medical staff were asked several questions on the pupillary examination. Reliability of PLD ratio were assessed via standard error of mean (S.E.M) of PLD ratios for 3 different subjects each imaged under constant illumination and fixation but from different angles to the optical axis.Results:Analysis of questionnaire data together with PLD ratios revealed that ~ 14% and 10% of participants could estimate the pupillary size in darkness and bright light respectively but none were simultaneously accurate indicating that subjective assessment of pupillary size was unreliable. The approach towards a systematic pupillary examination was inconsistent among the participants. The PLD ratio was found to be a reliable measure of pupillary size with standard error of mean below 0.1 mm for the three subjects tested.Conclusion:Static pupillary sizes can be objectively and consistently evaluated using PLD ratios using a two-box method. PLD ratios are resistant, within limits, to changes in imaging angle or choice of para-horizontal axes for measurement.

Purpose To characterize effects of muscarinic antagonist, atropine (A), and α-adrenergic agonist, phenylephrine (P), on mydriasis and light-evoked signaling in mice anesthetized by ketamine and xylazine (K+X). Methods Pupillary areas of anesthetized C57BL/6 mice were measured with or without topical application of either A or A+P. Dark-adapted ERGs were recorded from 2–4 month old C57BL/6 and 7.5 month old albino hrhoG/hrhoG mice following application of A or P singly or in combination before or after induction of K+X anesthesia. Effects of GABA were tested in the hrhoG/hrhoG mice. Results K+X anesthesia resulted in maximal mydriasis that was not enhanced by A or A + P. Dark-adapted b-wave amplitudes (−1.3 log sc td s) after K+X anesthesia were similar with or without A or P. A+P in the presence of K+X produced a slow growth in b-wave amplitude, reaching a plateau of two-fold enhancement in 1 h. Recordings with varying flash-energies revealed that the effects of A + P were on the maximum amplitude of the a- and b-waves and not on their sensitivity. Scotopic threshold responses were augmented as well. In photoreceptor-degenerated mice (hrhoG/hrhoG), an electronegative ERG wave recorded with K + X or K + X + A, was converted to a GABA-sensitive response with two electropositive components with A+P after K + X. Conclusion Topical administration of A, and P together, but not separately in presence of K+X, leads to a slow, dramatic enhancement of a- and b-waves by an unknown mechanism independent of pupil dilatation.

All three classes of receptors for the inhibitory neurotransmitter GABA (GABAR) are expressed in the retina. This study investigated roles of GABAR, especially GABACR (GABA(A)-ρ), in retinal signaling in vivo by studying effects on the mouse electroretinogram (ERG) of genetic deletion of GABACR versus pharmacological blockade using receptor antagonists. Brief full-field flash ERGs were recorded from anesthetized GABACR−/− mice, and WT C57BL/6 (B6) mice, before and after intravitreal injection of GABACR antagonists, TPMPA, 3-APMPA, or the more recently developed 2-AEMP; GABAAR antagonist, SR95531; GABABR antagonist, CGP, and agonist, baclofen. Intravitreal injections of TPMPA and SR95531 were also made in Brown Norway rats. The effect of 2-AEMP on GABA- induced current was tested directly in isolated rat rod bipolar cells, and 2-AEMP was found to preferentially block GABACR in those cells. Maximum amplitudes of dark (DA) and light-adapted (LA) ERG b-waves were reduced in GABACR−/− mice, compared to B6 mice, by 30–60%; a-waves were unaltered and oscillatory potential amplitudes were increased. In B6 mice, after injection of TPMPA (also in rats), 3-APMPA or 2-AEMP, ERGs became similar to ERGs of GABACR−/− mice. Blockade of GABAARs and GABABRs, or agonism of GABABRs did not alter B6 DA b-wave amplitude. The negative scotopic threshold response (nSTR) was slightly less sensitive in GABACR−/− than in B6 mice, and unaltered by 2-AEMP. However, amplitudes of nSTR and photopic negative response (PhNR), both of which originate from inner retina, were enhanced by TPMPA and 3-APMPA, each of which has GABAB agonist properties, and further increased by baclofen. The finding that genetic deletion of GABACR, the GABACR antagonist 2-AEMP, and other antagonists all reduced ERG b-wave amplitude, supports a role for GABACR in determining the maximum response amplitude of bipolar cells contributing to the b-wave. GABACR antagonists differed in their effects on nSTR and PhNR; antagonists with GABAB agonist properties enhanced light-driven responses whereas 2-AEMP did not.