GABAa and GABAc receptor mediated influences on the intensity-response functions of the b- and d-wave in the frog ERG.

We assessed the contribution of GABAa and GABAc receptors to GABAergic effects on b- and d-wave in frog ERG in a wide range of light stimulation conditions. The amplitude of both b- and d-wave was increased during GABAa receptor blockade by bicuculline as well as during additional GABAc receptor blockade by picrotoxin. The effects of GABAa receptor blockade were more pronounced in light adaptation conditions. They strongly depended on stimulus intensity and showed considerable ON/OFF-response asymmetry. The effects of GABAc receptor blockade were more pronounced in dark adaptation conditions. They didn't vary much with stimulus intensity and showed little ON/OFF-response asymmetry.

Effect of GABAergic blockade on light responses of frog retinal ganglion cells.

The effect of GABAergic blockade by picrotoxin on ganglion cells (GC) activity was investigated in perfused dark adapted eyecups of frog (Rana ridibunda). PT had diverse effects on the light responses of GC in contrast to its uniform potentiating effect on the amplitude of the ERG b- and d-wave. In some (n=32) of PT-sensitive ON-OFF GC the ON and OFF responses were changed in a similar manner (both responses were potentiated or both were inhibited), but in the other (n=10) the both responses were changed in a different manner. PT influenced differentially the activity of OFF GC (n=17) as well. It not only potentiated or inhibited their light responses, but changed also the temporal characteristics of the responses. Some tonic cells became phasic ones and in some phasic cells a late component appeared under the influence of PT. In some cases (n=4) the GABAergic blockade changed the apparent cell's type, because of appearance of a new type of response (ON or OFF) non-existing before the blockade. Our results indicate that the GABAergic interneurons are involved in different networks in the inner plexiform layer of frog retina.

Participation of the GABAergic system in the action of 2-amino-4-phosphonobutyrate on the OFF responses of frog retinal ganglion cells.

Perfusion of dark adapted frog eyecups with the ON pathway blocker 2-amino-4-phosphonobutyrate (APB) not only abolished the ganglion cells (GCs)' ON responses and the ERG b-wave, but it markedly potentiated the OFF responses of all ON-OFF and phasic OFF GCs and the d-wave amplitude of a simultaneously recorded ERG as well. The blockade of GABA(A) and GABA(C) receptors by picrotoxin eliminated this potentiating effect in 24 out of 41 GCs, although in the rest of the cells it did not produce any change in the APB effect. On the other hand, the d-wave potentiation was preserved during the GABAergic blockade in all experiments. Our results indicate that GABAergic transmission is involved in the inhibition exerted by the ON upon the OFF channel in part of the ON-OFF and phasic OFF GCs in the frog retina. The tonic OFF GCs probably do not receive an inhibitory input from the ON channel, because their light responses were not altered either by APB alone or by APB during blockade of GABA(A) and GABA(C) receptors.

Immunocytochemical and electrophysiological characterization of GABA receptors in the frog and turtle retina.

The expression of GABA receptors (GABARs) was studied in frog and turtle retinae. Using immunocytochemical methods, GABA(A)Rs and GABA(C)Rs were preferentially localized to the inner plexiform layer (IPL). Label in the IPL was punctate indicating a synaptic clustering of GABARs. Distinct, but weaker label was also present in the outer plexiform layer. GABA(A)R and GABA(C)R mediated effects were studied by recording electroretinograms (ERGs) and by the application of specific antagonists. Bicuculline, the GABA(A)R antagonist, produced a significant increase of the ERG. Picrotoxin, when co-applied with saturating doses of bicuculline, caused a further increase of the ERG due to blocking of GABA(C)Rs. The putative GABA(C)R antagonist Imidazole-4-acidic acid (I4AA) failed to antagonize GABA(C)R mediated inhibition and, in contrast, appeared rather as an agonist of GABARs.

Effect of 2-amino-4-phosphonobutyrate on the OFF responses of frog retinal ganglion cells and local ERG after glycinergic blockade.

Perfusion with the ON channel blocker 2-amino-4-phosphonobutyrate (APB) of dark adapted frog eyecups not only abolished the ganglion cells' (GC) ON responses and the ERG b-wave, but markedly potentiated the OFF responses of ON-OFF and phasic OFF-GCs and the d-wave amplitude of simultaneously recorded local ERG. Glycinergic blockade by strychnine prevented this potentiating effect in 31 out of 69 GCs, but did not change it at all in the other cells. At the same time the d-wave potentiation was preserved during the glycinergic blockade in all eyecups. The results indicate that glycinergic transmission is involved in the inhibition exerted from ON upon OFF channel in some but not all frog retinal GCs.

Influence of picrotoxin and strychnine on the spectral sensitivity of the turtle ERG b- and d-wave: I. Dark adaptation.

The ERG ON- (b-wave) and OFF-response (d-wave) to differently coloured stimuli was studied using a wide range of stimulus intensities in dark adapted turtle retina. The intensity-response curve of the b-wave showed saturation but that of the d-wave, decline in the high-intensity stimulus range. The curves of the relative spectral sensitivity of the ERG ON- and OFF-response were similar and showed a maximum in the longwave part of the spectrum. GABAergic blockade by 50 mumol L-1 picrotoxin caused an increase of the sensitivity, contrast sensitivity and the amplitude range of both ON- and OFF-responses without narrowing of the response dynamic range. In the range of lower stimulus intensities the ON-responses to blue stimuli and the OFF-responses to red stimuli were affected to a greater extent. An increased ERG b- and d-wave sensitivity was also observed during glycinergic blockade by 50 mumol L-1 strychnine. In the low intensity stimulus range the effect was maximal on the ON-response to blue stimuli and on the OFF-responses to 570 nm stimuli. It was concluded that the GABA- and glycinergic systems in the retina equalize rather than make different the relative spectral sensitivities of the ON- and OFF-responses.

Influence of picrotoxin and strychnine on the spectral sensitivity of the turtle ERG b- and d-waves: II. Light adaptation.

The aim of the present work was to investigate the role of GABA and glycine, the two main inhibitory neurotransmitters in the retina, in the spectral sensitivity coding under conditions of light adaptation. To study this question, spectral sensitivity curves, based on the turtle ERG responses to stimuli of different wavelengths, were constructed. The spectral sensitivity curves. obtained before and during treatment with picrotoxin (PT), a GABAA antagonist, or with strychnine (ST), a glycine antagonist, were compared. Both PT and ST increased the b- and d-wave absolute sensitivity in a wavelength-dependent manner. PT significantly changed the shape of the b- and d-wave spectral sensitivity curves and the latter lost their peaks. It is concluded that, under conditions of light adaptation, GABA and glycine took part in the formation of the b- and d-wave spectral sensitivity curves, that both of them exerted an effect on the gain and that furthermore,GABA had a well pronounced effect on the tuning of the spectral sensitivity curves.

The application of in vitro isolation, cryopreservation and patch-clamp microelectrode recording methods to adult rat thymic nurse cells.

Characterisation of the ionic mechanisms possessed by immune cells has begun to reveal a range of transmembrane ion channel properties which may have immunological significance. Since thymic epithelial cells appear to influence selection of the T cell repertoire, understanding their membrane physiology may be of importance. A method is therefore outlined for the targeting of patch-clamp electrophysiological measurements to acutely isolated, fresh and cryopreserved adult rat thymic nurse cells. These cells represent a discrete and predominantly cortical population of epithelium. Cells were separated by enzymatic and mechanical dispersal of thymus, enriched by sedimentation and identified on the basis of their characteristic lympho-epithelial, multi-cellular morphologies. Phase-bright cells retaining this anatomical form survived freezing and the voltage-gated conductances in such cells are described. The merits of this approach include the preparation and storage of mature differentiated phenotypes of immune cells for in vitro studies. Their preservation permits temporal separation of experiments, correlative experimentation on identical samples, optimises cell preparation in terms of cost and labour, and reduces animal and tissue requirements.

ERG OFF response in frog retina: light adaptation and effect of 2-amino-4-phosphonobutyrate.

The intensity-response (V/log I) function of ERG OFF response (d-wave) in dark and light adapted superfused frog eyecups was investigated before and after blockade of the retinal ON channel by 2-amino-4-phosphonobutyrate (APB). The V/log I function of the dark adapted d-wave had two distinct components, each of them consisting of an ascendent and descendent part. In eyes adapted to mesopic or photopic background the V/log I function had only one component. It was shifted to the right along the intensity axis, had a steeper slope and a higher maximal response amplitude compared with the two components of the dark-adapted V/log I curve. Perfusion with 200 mumol APB markedly increased the d-wave amplitude at all stimulus intensities except for the threshold ones in both dark and light adapted eyes. The position of the V/log I curve was shifted slightly to the left along the intensity axis in dark adapted eyes, but was not changed in light adapted eyes. Thus the adaptational mechanism responsible for changes in the decremental sensitivity with increased background illumination was not altered by APB. The effect of APB was studied also in chromatically adapted eyes, in which the responses were predominantly mediated by one photoreceptor type. The results showed that the potentiating effect of APB on d-wave did not depend on photoreceptor input.

Effect of 2-amino-4-phosphonobutyrate on ERG OFF-response after glycinergic and GABAergic blockade.

Superfusion with 200 microM 2-amino-4-phosphonobutyrate (APB) of dark and chromatically adapted frog eyecups caused marked potentiation of the ERG OFF-response (d-wave). Blockade of the glycinergic synapses by strychnine did not change this effect at all. Blockade of the GABAergic synapses by picrotoxin slightly diminished the effect of APB in chromatically-adapted eyes with isolated cones' activity, and did not change it in dark-adapted eyes. The results indicate that the action of APB on ERG OFF-response does not depend significantly on GABAergic and glycinergic neurotransmission in frog retina.

Comparative investigation of retinal responses to brief light stimuli: 2-amino-4-phosphonobutyrate studies--I. Frog retina, Rana ridibunda.

1. Electroretinogram (ERG) and responses of single ganglion cells to 75 microseconds light flashes, applied at two different backgrounds, were studied. Additionally, a stimulation with long-lasting stimuli (ordinarily 5 sec ON-, 5 sec OFF-) was used. Both white and coloured light stimuli were presented. 2. 150 microM 2-amino-4-phosphonobutyrate (APB) was used to separate OFF- from ON- channels. 3. Before APB application, one or two components in the impulse activity in response to a flash were observed, depending on the type of ganglion cell (ON-, OFF- or ON-OFF). the latency of the first component was 60 msec and the latency of the second one was from 160 to 430 msec on average, at different background conditions. APB abolished the first component and enhanced the second one. 4. By means of APB, two components were shown to exist in the main positive wave of the flash ERG. APB abolished the first component and did not influence or enhance the second one. 5. The data obtained show that both ON- and OFF- channels take part in the generation of the frog retinal responses to brief stimuli.

Comparative investigation of retinal responses to brief light stimuli: 2-amino-4-phosphonobutyrate studies--II. Turtle retina (Emys orbicularis).

1. Electroretinogram (ERG) and responses of single ganglion cells to 75 microseconds light flashes, presented on two different backgrounds, were studied. Additionally, stimulation with long lasting stimuli (ordinarily 5 sec ON-, 15 sec OFF-) was used. 2. 2-amino-4-phosphonobutyrate (APB) at a concentration of 450 microM on average was used to separate OFF- from ON- channels. It is known, that in other species APB selectively blocks the activity of ON- channel only. 3. The existence of APB- sensitive membrane receptors was demonstrated in the turtle retina. As in other species, APB abolished the ERG b-wave and enhanced the d-wave, when long lasting stimulation was used. 4. By means of APB, two components were shown to exist in the ERG positive wave in response to a flash. APB abolished the first component and did not influence or enhanced the second one. 5. By means of APB, one or two components in the impulse activity in response to flash were demonstrated, depending on the type of ganglion cell (ON-, OFF- or ON-OFF). The latency of the first component was 70 msec and the latency of the second one 210 msec on average. APB abolished the first component, and enhanced the second one. 6. The data obtained show that both ON- and OFF- channels take part in the generation of the turtle retinal responses to brief stimuli. 7. Based on the results obtained, some peculiarities of the network organization of the ganglion cells' receptive fields are discussed.

Participation of the GABAergic system of the turtle retina in the light adaptation process.

A comparative study was made of the ERG b- and d-wave intensity-response functions before and after GABAergic blockade by means of 0.4 mmol l-1 picrotoxin. A wide range of background intensities, including part of the high photopic range, were used. The intensity-response functions of both the ERG waves fitted well to the Michaelis-Menten equation (V/Vmax = In/(In + sigma n). A sigma-value decrease and a Vmax and n increase were observed after picrotoxin treatment. The analysis of the intensity-response functions shows that, under a wide range of backgrounds, the GABAergic neurons influence the ON- and OFF-response in the distal retina in a similar way. They decrease the gain and the contrast gain of the b- and d-wave generating neuronal mechanisms and widen the intensity span of their responses under given background illumination. The GABAergic system is involved also in response 'scaling' in the ON- and OFF-channels in the distal retina. A very important effect in this respect seems to be the equalization of the range of the responses to increment stimuli under different backgrounds.

Comparative studies on the effects of some enkephalin agonists on the ERG of frog and turtle.

1. Effects of mu-agonist morphine in a concentration of 1.10(-5)-1.10(-6) M and delta-agonist Dalargin in a concentration of 1.10(-5)-1.10(-8) M on electroretinogram (ERG) of frog (mixed type of retina) and turtle (predominantly cone retina) were investigated. 2. The enkephalin agonists studied influenced both types of retina in a different manner, producing mainly inhibitory effects. 3. Variability in mu-receptors sensitivity in the turtle retina, most probably depending on the seasons, was observed. 4. The data obtained show that opiate-sensitive mechanisms control the bioelectric activity of the distal layers of frog and turtle retina. 5. Species specificity and variability in sensitivity of the opiate receptors are discussed.

Influence of picrotoxin on the retinal sensitivity at different levels of background illumination.

The intensity-response function of the b-wave of the ERG in the retina of the turtle (Emys orbicularis) was investigated at two different background illuminations--0.01 and 10 lx, before and after blockade of the GABA-ergic transmission by 0.4 mmol/l picrotoxin (PT). A shift of the curve to the left along the intensity axis at both backgrounds was observed after PT treatment. The region where the b-wave amplitude was linear function of the logarithm of the stimulus intensity was also considerably narrowed. The registered experimental data fitted in well with the Naka-Rushton equation. The participation of GABA-ergic neurones in the processes of light adaptation and gain control in the retina is discussed.

Participation of GABA in the sensitivity control of the OFF-response in the turtle ERG.

The intensity-response--V/log I, function of the d-wave in the turtle ERG was investigated before and after blockade of the GABAergic transmission by 0.4 mmol/l picrotoxin (PT) under conditions of two adapting background illuminations--0.01 and 10 lx. After PT the d-wave V/log I curves showed a shift to the left along the intensity axis under both adapting conditions, revealing an increase in the gain and the contrast sensitivity of the retinal elements involved in the d-wave generation. These changes were similar to the changes in the b-wave V/log I function described under similar conditions. The relative increase of the b- and d- wave amplitude had approximately equal values at both background levels at higher stimulus intensities. In the range of low stimulus intensity, however, at background illumination of 0.01 lx the relative increase of the d-wave was considerably greater than that of the b-wave. The experimental data obtained were best fitted by the Naka-Rushton equation. The participation of the GABAergic neurons on the d-wave sensitivity control is discussed.

Effects of picrotoxin on the turtle's electroretinogram under different background illumination.

The effect of 0.4 mM solution of picrotoxin (PT) on the electroretinogram (ERG) of isolated eyecup preparations of turtle (Emys orbicularis) was investigated under two different backgrounds (Ib)--scotopic (0.009 lx) and mesopic (9 lx). The test stimulus intensity was 46 lx. An increase of the amplitudes of all ERG waves was observed after PT application. The relative increase of the d-wave was the most pronounced one and was similar under the two Ib. The relative increase of the b-wave, however, was greater in mesopic than in scotopic conditions. The two subcomponents of the a-wave were differentially influenced by PT. The development of a new steady level of b-wave sensitivity was delayed after PT.

On the mechanisms underlying appearance of responses to movement, directional sensitivity and velocity tuning of the cat's striate cortical neurons.

The responses to moving and stationary stimuli of 27 cat's striate cortical units were studied. Two stationary light bars located in different parts of the receptive field were used. The order of presentation and the time-interval between the stimuli varied; so, the presentation of a pair of stationary stimuli was an analogue of a moving stimulus. It was shown that responses occurred in neurons previously unresponsive to stationary stimuli when two stationary stimuli were presented successively in certain order. In the direction-sensitive units an asymmetry of the temporal course of the inhibitory processes was observed. The inhibitory zone located on the side of the preferred direction of movement was characterized by an early inhibitory phase followed by a phase of disinhibition and by a second inhibitory phase. For the inhibitory zone located on the side of the null direction no disinhibitory phase was demonstrated. The significance of the spatial and temporal characteristics of the receptive field for the appearance of responses to movement, the directional sensitivity and the velocity tuning in striate neurons is discussed.

[Mechanism of responses to movement in the visual cortex neurons unresponsive to light flicker]. / O mekhanizme vozniknoveniia otvetov na dvizheniia u neironov zritel'noi kory, ne otvechaiushchikh na svetovye mel'kaniia.

The responses to moving and stationary stimuli in cat's striate cortical units were studied. Two stationary light bars, one located in the receptive field centre, and another--in the periphery, were used. The sequence of presentation and the time-interval between the stimuli varied thus making the presentation of a pair of stationary bars an analogue of a moving stimulus. Responses occurred in the neurons previously unresponsive to stationary stimuli when two stationary stimuli were presented successively in certain order. A model is proposed explaining the occurrence of responses to moving stimuli in the neurons, unresponsive to stationary ones.