Biomedical subjects

J Ammermüller

Publications and source records attributed to J Ammermüller.

At least 19 recordsLinked to original sources

PubMed2001

Visual transmission deficits in mice with targeted disruption of the gap junction gene connexin36.

In the mammalian retina, rods feed into the cone pathway through electrotonic coupling, and recent histological data suggest the involvement of connexin36 (Cx36) in this pathway. We therefore generated Cx36 null mice and monitored the functional consequences of this deficiency on early visual transmission. The homozygous mutant mice had a normally developed retina and showed no changes in the cellular organization of the rod pathway. In contrast, the functional coupling between AII amacrine cells and bipolar cells was impaired. Recordings of electroretinograms revealed a significant decrease of the scotopic b-wave in mutant animals and an increased cone threshold that is compatible with a distorted, gap junctional transmission between AII amacrine cells and cone bipolar cells. Recordings of visual evoked potentials showed extended latency in mutant mice but unaffected ON and OFF components. Our results demonstrate that Cx36-containing gap junctions are essential for normal synaptic transmission within the rod pathway.

Animals↗

PubMed2001

Population coding of motion patterns in the early visual system.

Using extracellular recordings and computational modeling, we study the responses of a population of turtle (Pseudemys scripta elegans) retinal ganglion cells to different motion patterns. The onset of motion of a bright bar is signaled by a rise of the population activity that occurs within less than 100 ms. Correspondingly, more complex stimulus movement patterns are reflected by rapid variations of the firing rate of the retinal ganglion cell population. This behavior is reproduced by a computational model that generates ganglion cell activity from the spatio-temporal stimulus pattern using a Wiener model complemented by a non-linear contrast gain control feedback loop responsible for the sharp transients in response to motion onset. This study demonstrates that contrast gain control strongly influences the temporal course of retinal population activity, and thereby plays a major role in the formation of a population code for stimulus movement patterns.

Animals↗

PubMed2000

Population coding in spike trains of simultaneously recorded retinal ganglion cells.

To achieve a better understanding of the parallel information processing that takes place in the nervous system, many researchers have recently begun to use multielectrode techniques to obtain high spatial- and temporal-resolution recordings of the firing patterns of neural ensembles. Apart from the complexities of acquiring and storing single unit responses from large numbers of neurons, the multielectrode technique has provided new challenges in the analysis of the responses from many simultaneously recorded neurons. This paper provides insights into the problem of coding/decoding of retinal images by ensembles of retinal ganglion cells. We have simultaneously recorded the responses of 15 ganglion cells to visual stimuli of various intensities and wavelengths and analyzed the data using discriminant analysis. Models of stimulus encoding were generated and discriminant analysis used to estimate the wavelength and intensity of the stimuli. We find that the ganglion cells we have recorded from are non-redundant encoders of these stimulus features. While single ganglion cells are poor classifiers of the stimulus parameters, examination of the responses of only a few ganglion cells greatly enhances our ability to specify the stimulus wavelength and intensity. Of the parameters studied, we find that the rate of firing of the ganglion cells provides the most information about these stimulus parameters, while the timing of the first action potential provides almost as much information. While we are not suggesting that the brain is using these variables, our results show how a population of sensory neurons can encode stimulus features and suggest that the brain could potentially deduce reliable information about stimulus features from response patterns of retinal ganglion cell populations.

Action Potentials↗

PubMed1998

UV-sensitive input to horizontal cells in the turtle retina.

Microspectrophotometry, electroretinography and behavioural studies have indicated that ultraviolet (UV) light contributes to functional vision in various vertebrate species. Based on behavioural evidence, this was also suggested for turtle vision. In order to reveal the interactions underlying detection of UV light in the distal retina, we recorded intracellularly the photoresponses of cones and horizontal cells in retinas of Pseudemys scripta elegans and Mauremys caspica and calculated the action spectra of these cells under different conditions of adaptation. In the dark-adapted retina, all three types of horizontal cells; luminosity-type, red/green chromaticity-type and yellow/blue chromaticity-type exhibited increased sensitivity in the UV region of the spectrum. However, chromatic adaptation indicated that only the yellow/blue chromaticity-type horizontal cells received excitatory input from UV-sensitive cones with peak sensitivity approximately 360 nm. The enhanced UV sensitivity of luminosity-type horizontal cells probably reflected the beta-band of the long-wavelength sensitive visual pigment as indicated by the action spectra of dark-adapted L-cones. It is suggested that the enhanced UV sensitivity of red/green chromaticity-type horizontal cells reflects the beta-band of the medium-wavelength sensitive visual pigment. Transmission measurements of the optical media (cornea, lens and vitreous) indicated that UV vision can be functional under normal circumstances.

Adaptation, Physiological↗

PubMed1997

Synaptic inputs to identified color-coded amacrine and ganglion cells in the turtle retina.

Previous studies have proposed models of the specific synaptic circuitry responsible for color processing in the turtle retina. To determine the accuracy of these models of the circuits underlying color opponency in the inner retina of the turtle (Pseudemys scripta), we have studied the physiology, morphology, and synaptic connectivity of identified amacrine and ganglion cells. These cells were first characterized electrophysiologically and were then stained with horseradish peroxidase. Postembedding electron immunocytochemistry for gamma-aminobutyric acid (GABA) and glycine was used to reveal the neurochemical identity of their synaptic inputs. The red-ON/green, blue-OFF small-field ganglion cell, classified as G24, branched primarily in strata S1, S4, and S5 of the inner plexiform layer (IPL). Ganglion cell G24 showed a complex receptive field organized into a red-ON center surrounded by an inhibitory region, which, in turn, was surrounded by a second excitatory region. Only the center responses were color opponent. The red-OFF/green, blue-ON large-field, stellate amacrine cell, classified as A23b, stratified exclusively in stratum S2, near the S2/S3 border. The color-coded center was surrounded by a luminosity, red-sensitive surround. Synaptic input to G24 and A23b was dominated by amacrine cells (89% and 87%, respectively). G24 received significant input from amacrine cell profiles with GABA (13% of total) as well as glycine (11% of total) immunoreactivity, mostly in the proximal stratum S5 of the IPL (64% and 67% of the total GABA- and glycine-immunoreactive input, respectively). Bipolar cell synaptic input was also found predominantly in S4 and S5 (89%). In contrast, we found no glycine-immunoreactive input to A23b, and the density of the GABA-immunoreactive amacrine cell synaptic input revealed a central (15%) to peripheral (3%) gradient within the dendritic tree. The results of the present study support the previous models of the synaptic circuitry responsible for color-opponent signal processing in the inner retina of the turtle.

Animals↗

PubMed1997

Neural circuitry and light responses of the dopamine amacrine cell of the turtle retina.

PURPOSE: To understand the circuitry and electrophysiology of the dopamine cells in the turtle retina. METHODS: Preembedding immunocytochemistry for tyrosine hydroxylase (Toh) was done on vibratome sections of turtle retina. Resultant Toh-immunoreactive (Toh-IR) amacrine cells were then serially thin-sectioned for analysis by electron microscopy (EM). Some sections of Toh-IR cells also were post-embedding immunostained for glycine and GABA content. Intracellular recordings and dye markings were made from the turtle eyecup and slice preparation to determine the light responses of cells called A28, which have the same morphology as Toh-IR amacrine cells. RESULTS: Physiologically A28 cells were L-type (luminosity) and gave sustained depolarizing (ON-center) responses to light pulses. High intensity light pulses produced immediate transients and long depolarizations, lasting beyond the stimulus duration. An after-hyperpolarization and an antagonistic surround could be elicited. EM reconstruction of a Toh-IR cell revealed new circuitry over that described before (Pollard, J. & Eldred, W.D. (1990). J. Neurocytol. 19, 53-66). Bipolar ribbon synapses occurred in all three dendritic tiers. However, amacrine cell inputs dominated numerically (95% amacrine input, 5% bipolar input) many of them in a serial synaptic configuration. GABA+ inputs were seen but not glycine+ inputs. Output from Toh-IR profiles was primarily to large ganglion cell dendrites but also to bipolar cell axons, GABA-IR amacrines, unspecified amacrine cells and other Toh-IR dendrites. CONCLUSIONS: The synaptology of the dopamine cells of the turtle retina suggests that sustained inhibitory amacrine cell pathways, including GABAergic pathways, are chiefly responsible for their response characteristics at low light levels. Conversely, at higher light intensities, transient excitatory amacrine cells probably have influence.

Animals↗

PubMed1996

Quantitative anatomy, synaptic connectivity and physiology of amacrine cells with glucagon-like immunoreactivity in the turtle retina.

Although a wide variety of neuropeptides have been localized in vertebrate retinas, many questions remain about the function of these peptides and the amacrine cells that contain them. This is because many of these peptidergic amacrine cells have been studied using only immunocylochemical techniques. To address this limitation, the present study used a combination of quantitative anatomy, biochemistry and electrophysiology to examine amacrine cells in the turtle retina that contain the neuropeptide glucagon. In the turtle retina, there is a small population of 2500 glucagonergic amacrine cells, which probably represents < 1% of the total number of amacrine cells. Circular distribution statistics indicated that many of these tristratified amacrine cells had asymmetric dendritic arborizations that were radially oriented toward the retinal periphery. The cells were found to have similar dendritic coverage factors, to be distributed in a non-random arrangement in all regions of the retina, and to peak in density in the visual streak region. Electron microscopic studies indicated that glucagonergic amacrine cells made synaptic contacts primarily with other amacrine cells, and small numbers of bipolar cells. The synaptic inputs and outputs were balanced in the inner strata of the inner plexiform layer, and were biased toward synaptic outputs in the outer strata of the inner plexiform layer. These contacts involved small unlabelled synaptic vesicles, and not the large labelled dense core vesicles also found in these neurons. The biochemical studies indicated that glucagon could be released from the retina in a calcium dependent manner by high potassium stimulation. The electrophysiology found no color opponency, and the glucagonergic amacrine cells gave sustained hyperpolarizing responses to small stimulation spots and had antagonistic surrounds. The results of these studies suggest that there are significant regional specializations of glucagonergic amacrine cells, and that they may provide OFF-modulation in interactions between the ON-and OFF-centre visual pathways in the turtle retina.

Animals↗

PubMed1996

Effects of horizontal cell network architecture on signal spread in the turtle outer retina. Experiments and simulations.

In the Pseudemys turtle retina five functionally distinct, electrically coupled networks of horizontal cells distribute signals in the outer plexiform layer. These networks differ significantly in their architecture, as determined by intracellular labeling with Neurobiotin after physiological recording and identification. The density of H1 horizontal cells is highest, ranging around 1800 cells/mm2 at approximately 2.3 mm eccentricity. H1 horizontal cell somata are connected via 6-10 thin, short dendrites. The H1 horizontal cell axon terminal network is composed of thick axon terminals, forming a three-dimensional, sheath-like structure. Networks of coupled H2 and H3 horizontal cells have cell densities of around 210 cells/mm2 and 350 cells/mm2, respectively, at the same eccentricity of 2.3 mm. Cell bodies are connected with 6-12 long, thin dendrites. Here we report for the first time H4 horizontal cell networks. Cell density is approximately 970 cells/mm2 at 2 mm eccentricity, and cell bodies are connected with 6-10 thin, short dendrites. General properties of passive voltage spread were compared for three of these horizontal cell networks using NeuronC. Realistic network architectures were obtained by digitizing the intracellularly labeled networks, respectively. One network obtained from coupled H1 horizontal cell bodies, one from coupled H1 horizontal cell axon terminals, and one from H2 horizontal cells were simulated. These three realistic networks were compared with an artificial, electrically coupled regular triangular network. Passive signal spread in these networks strongly depended on the exact network architecture using otherwise identical parameters. Changes in coupling strength affected signal spread in these networks differently. As in the experimental situation, changes in synaptic conductance influenced signal spread. Some principal effects of extensively coupled horizontal cells on photoreceptor signal processing were simulated with one type of photoreceptor connected by telodendria, synapsing onto an underlying triangular network and receiving feedback synapses. Under certain conditions, spatial information is coded in single photoreceptors. This was also the case in the experimental situation. In the simulation, spatial filter adjustment for optimal spatial coding in photoreceptors can be achieved by changing coupling strength in the horizontal cell network.

Animals↗

PubMed1995

The organization of the turtle inner retina. I. ON- and OFF-center pathways.

Intracellular recordings and dye injections of Lucifer yellow, horseradish peroxidase, or Neurobiotin were made in bipolar, amacrine, and ganglion cells of the Pseudemys turtle retina. By using a standard light-stimulation protocol in a sample of 375 labeled neurons, we were able to identify morphological and physiological characteristics of 11 types of bipolar cell, 37 types of amacrine cell, and 24 types of ganglion cell. To make sense of these data, we have chosen to group the 72 essentially different neuron types into traditional, functionally significant pathways. In this paper we look at the neuronal types in the inner plexiform layer (IPL) in terms of their contribution to generalized luminosity responses such as sustained ON- or OFF-center and transient ON-OFF ganglion cells; in the companion paper (J. Ammermüller, J.F. Muller, and H. Kolb, 1995, J. Comp. Neurol. 358:35-62) we look at them in terms of their involvement in color opponency and directional selectivity. A functional organization of the turtle IPL into OFF sublaminae (strata 1 and 2) and ON sublaminae (strata 3, 4, and 5), as has been described for other vertebrate retinas, was quite clear for two varieties of OFF-center bipolar cells (B4 and B5) and for all four types of sustained ON-center bipolar cell (B1, B2, B6, and B7). Thus, we found no sustained ON-center bipolar cell terminating in strata 1 and 2. We did, however, see three varieties of sustained OFF-center bipolar cells (B3, B9, and B10) having axon terminals in strata 3-5 (the ON sublamina) in addition to their terminations in stratum 1 or 2 (the OFF sublamina). Monostratified sustained ON- and OFF-center amacrine and ganglion cells rigidly obeyed the border of ON and OFF sublaminae. However, multistratified and diffuse sustained amacrine and ganglion cells could be either ON-center or OFF-center, and they did not strictly obey the border: such ON-center cells always had processes in one of the ON sublaminae (strata 3-5), and the equivalent OFF-center cells always had processes in one of the OFF sublaminae (strata 1 and 2). Monostratified transient amacrine and ganglion cells were concentrated in the middle of the IPL (around stratum 3), whereas bi-, tri-, or multistratified transient amacrine or ganglion cells always had processes in both the ON and the OFF sublaminae.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1995

The organization of the turtle inner retina. II. Analysis of color-coded and directionally selective cells.

Color coding and directional selectivity (DS) of retinal neurons were studied in the Pseudemys turtle by using similar intracellular recording and staining techniques as in the preceding paper (J. Ammermüller and H. Kolb, 1995, J. Comp. Neuronal. 358:1-34). Color-coded responses were elicited by red (621 or 694 nm), green (525 or 514 nm), and blue (455 nm) light flashes. In addition to red/green and yellow/blue types of chromaticity horizontal cells, in our sample of 305 identified cells we found that 17% of bipolar cells, 6.5% of amacrine cells, and 18% of ganglion cells exhibit color-coded responses. DS responses were found in 37% of the tested ganglion cells and 41% of the tested amacrine cells. Two morphologically identified bipolar cell types, B10 and B11, were red-ON/blue-OFF and red-OFF/green, blue-ON, respectively. Of five identified amacrine cell types, three were red-OFF/blue-ON center (A1, A3, A23b), one was red-OFF/green-ON center (A32), and one (A33) was double color-opponent of red-ON/blue-OFF center:red-OFF/blue-ON surround. Five ganglion cell types had variously color-coded centers (G14 and G24) or surrounds (G3 and G18), including one type, G6, that was double color-opponent (red-OFF/green-ON center:red-ON/green-OFF surround). Responses to colors were found primarily in sustained responses of bipolar and ganglion cells. However, in amacrine cells, transient components of the response also showed color dependence. Red-OFF-center responses were found in ganglion cells that were in a position to make connections at the strata 2/3 border with the red-OFF bipolar cell (B11); red-ON-center responses occurred in ganglion cells with branches in stratum 4 of the IPL where the red-ON-center bipolar (B10) ended. Blue-ON-center signals appeared to be processed mainly in strata 1-2/3, and blue-OFF-center signals in strata 3-5 of the IPL, with contributions of amacrine cells and bipolar cells. Labeled DS amacrine cells could be identified as A9, A20, and A22, and ganglion cells as G19, G20, and G24. The latter type (G24) showed DS and color coding. All response types (ON-center, OFF-center, ON-OFF) were encountered. DS amacrine cells were monostratified near the middle of the IPL, whereas DS ganglion cells were mono-, bi-, and multistratified, although all DS ganglion cells had one feature in common: they had dendrites in stratum 1 of the IPL.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1995

Substance P: a neurotransmitter of amacrine and ganglion cells in the vertebrate retina.

A short history and summary of the occurrence of substance P in the vertebrate body is presented. Substance P is now generally accepted to be a neurotransmitter and can be visualized by immunocytochemistry to occur in various nerve cells in the CNS. In the retina, substance P-immunoreactivity (SP-IR) occurs in amacrine cell populations in all the species so far studied. In some vertebrates retinas SP is also apparent in one or more ganglion cell types. Anatomical investigations have revealed the morphology and connectivity of SP-IR amacrine cells: they branch in several strata of the inner plexiform layer receiving input from bipolar and amacrine cells and making synapses upon bipolar and ganglion cells. Most commonly SP-IR amacrines emit axon-like process that pass to both the outer plexiform layer and the ganglion cell and nerve fiber layers. These processes often end upon the retinal vasculature. SP-IR ganglion cells have been described in turtle, rabbit and human retinas. In turtle, intracellular dye injection has revealed the morphology of one type of SP-IR ganglion cell as being a large-field monostratified cell with a branches in the outer stratum of the inner plexiform layer. It may correspond to a "Dogiel cell" type. Intracellular investigation of SP-IR amacrine cells in turtle reveal their physiological responses to be ON-OFF in nature with some color-coding characteristics. In general SP acts as an excitatory neurotransmitter raising the spontaneous activity level of ganglion cell responses. The SP-IR ganglion cell is an OFF-center unit in the turtle retina and may be driven in the center of its receptive field by luminosity bipolar cells and in its surround by amacrine cells with color-opponent properties.

Animals↗

PubMed1994

Complexity and scaling properties of amacrine, ganglion, horizontal, and bipolar cells in the turtle retina.

In the present study we have evaluated the complexity and scaling properties of the morphology of retinal neurons using fractal dimension as a quantitative parameter. We examined a large number of cells from Pseudemys scripta and Mauremys caspica turtles that had been labeled using Golgi-impregnation techniques, intracellular injection of Lucifer Yellow followed by photooxidation, intracellular injection of rhodamine conjugated horseradish peroxidase, or intracellular injection of Lucifer Yellow or horseradish peroxidase alone. The fractal dimensions of two-dimensional projections of the cells were calculated using a box counting method. Discriminant analysis revealed fractal dimension to be a significant classification parameter among several other parameters typically used for placing turtle retinal neurons in different cell classes. The fractal dimension of amacrine cells was significantly correlated with dendritic field diameters, while the fractal dimensions of ganglion cells did not vary with dendritic field span. There were no significant differences between the same cell types in two different turtle species, or between the same types of neurons in the same species after labeling with different techniques. The application of fractal dimension, as a quantitative measure of complexity and scaling properties and as a classification criterion of neuronal types, appears to be useful and may have wide applicability to other parts of the central nervous system.

Animals↗

PubMed1994

Neurotensin-like immunoreactivity in locust supraesophageal ganglion and optic lobes.

A substance immunoreactive to antibodies directed against bovine neurotensin (NT) was localized in neurons in the supraesophageal ganglion (SEG) and optic lobes of larval and adult Locusta migratoria L. Two large somata were located in the caudal cortex, ventral to the calyces and symmetrical to the median of the SEG. Four smaller somata also in the caudal cortex were located as two symmetrical pairs at the level of the central body. These somata formed a diffuse network of varicose fibers from the superior lateral to the ventro-lateral protocerebrum between the pedunculi and frontal cortical region. Some fibers crossed the median to the contralateral sides of the SEG. Another pair of immunoreactive somata whose terminating processes remained unclear was found at the level of the antennal lobes. Intrinsic networks of fibers were labeled in the accessory medulla and in layer 4/5 of the medulla. These fibers originated from 8-10 small somata near the dorso-frontal rim of the medulla. All larval stages contained these NT-like immunoreactive structures. Results from isoelectric focusing and press-blot analysis of SEG homogenates, synthetic neurotensin and neurotensin fragments indicate that this substance is similar to bovine neurotensin(1-13).

Animals↗

PubMed1994

Receptive-field size of L1 horizontal cells in the turtle retina: effects of dopamine and background light.

1. The receptive-field size of turtle L1 horizontal cells was assessed qualitatively from the small-spot/full-field-response amplitude ratio. For quantitative evaluation, the length constant was derived from the response amplitude-spot radius relationship. 2. In each horizontal cell, the length constants were calculated for different intensities of the test light stimuli. The effects of dopamine and/or background light on the small-spot/full-field amplitude ratio and on the length constants were studied. 3. The receptive field of an L1 horizontal cell could not be defined by a single length constant of fixed value. Rather, the length constant changed with the experimental conditions. Two types of changes were noted. An instantaneous one, which was expressed in an increase in the length constant when the test flash was made brighter, and a slow one that occurred when the eyecup was exposed to dopamine. 4. Dopamine increased the small-spot/full-field amplitude ratio and reduced the length constant for a given full-field-response amplitude. It did not alter the responsiveness to light of the horizontal cells. These effects of dopamine were consistent with its action on the coupling resistance between adjacent horizontal cells. 5. Continuous background illumination increased the small-spot/full-field-response amplitude ratio whether studied in normal Ringer or during superfusion with dopamine solution. 6. The relationship between the length constants and the relative amplitude of the full-field responses did not change when the level of ambient illumination was raised either during superfusion with normal Ringer solution or during superfusion with dopamine solution. 7. These data indicate that background lights do not alter the receptive field size of turtle L1 horizontal cells.

Animals↗

PubMed1993

A geometrical description of horizontal cell networks in the turtle retina.

Networks of physiologically identified H2 horizontal cells in the turtle retina were labeled by intracellular injection of Neurobiotin. We obtained a quantitative description of the neighbourhood relations in the dye-coupled cell mosaics by using the somata as centers for the Voronoi-Delaunay construction. Computational models simulating the experimental data are presented.

Animals↗

PubMed1991

Synaptic inputs to physiologically defined turtle retinal ganglion cells.

Two physiologically distinct, HRP-marked turtle retinal ganglion cells were examined for their morphology, GABAergic, glycinergic, and bipolar cell synaptic inputs, using electron-microscopic autoradiography and postembedding immunocytochemistry. One cell was a color-opponent, transient ON/OFF ganglion cell. Its center response to red was a sustained hyperpolarization, and its center response to green was a depolarization with increased spiking at onset. The HRP-injected cell most resembled G6, from previous Golgi-impregnation studies (Kolb, 1982; Kolb et al., 1988). It was a narrow-field bistratified cell, whose two broad dendritic strata peaked at approximately levels L20-25 (sublamina a) and L60 (sublamina b) of the inner plexiform layer. Bipolar cell synapses onto G6 were found evenly distributed between its distal and proximal dendritic strata, spanning L20-75. These inputs probably originated from several different bipolar cells, reflecting the complexity of the center response. GABAergic inputs were found onto both the distal and proximal strata, from near L20-L85. Only a few glycinergic inputs, confined to dendrites at L50-70, were observed. A second ganglion cell type that we physiologically characterized and HRP-injected had sustained ON-center, sustained OFF-surround responses. Two examples were studied; both were bistratified in sublamina b, near L60-70 and L85-100, with branches up to near L40. They resembled G10, from previous Golgi-impregnation studies (Kolb, 1982; Kolb et al., 1988). One cell was partially reconstructed to look at the distributions of GABAergic and glycinergic amacrine cell, and bipolar cell inputs. Although synapses from bipolar cells were equally divided between the two major dendritic strata of G10, the inputs to the distal stratum were close to the soma, and the inputs to the more proximal stratum were on the peripheral dendrites. This arrangement may reflect input from two distinct types of ON-bipolar cell. GABAergic and glycinergic inputs to G10 costratified to both strata and to the distal branches; but where glycinergic inputs were found distributed throughout the arbor, GABAergic inputs appeared to be confined to peripheral dendrites. We hypothesize on the neural elements involved and the circuitry that may underlie the physiologically recorded receptive fields of these two very different ganglion cell types in the turtle retina.

Animals↗

PubMed1990

Evaluation of horseradish peroxidase as an intracellular stain in electrophysiological recording.

Iontophoresis of horseradish peroxidase into cells during intracellular electrophysiological recording to visualize the neurons recorded from is a powerful technique, but its success depends on a number of critical factors. The described agar injection technique is a simple method for the evaluation of several parameters at once, using procedures which closely resemble recording from, fixing, and processing to visualize the HRP in experimental tissue.

Electrophysiology↗

PubMed1990

A 'puff and advance' technique for visually controlled staining of turtle retinal ganglion cells.

We describe a 'puff and advance' technique for visually controlled staining of retinal ganglion cells (GCs) in the unfixed, living retina for light and electron microscopy. Glass microelectrodes are filled with rhodamine-isothiocyanate labeled horseradish peroxidase (Rh-HRP), or Lucifer yellow (LY), or a mixture of both, or with 5,6-carboxytetramethylrhodamine (5,6-Rh) and advanced tangentially through the GC layer with microscopic observation using epifluorescence. Brief "puffs" of LY or 5,6-Rh are constantly ejected from the advancing electrode tip by a train of negative current pulses. GC penetration is signaled by virtually instantaneous staining of its soma (and eventually its axon and dendrites if the electrode is not advanced further). An impaled GC can be electron densely stained with the Rh-HRP complex by switching to positive current pulses. The extent of dye filling is monitored through the microscope using a filter combination appropriate for the dye. After fixation, standard histochemical procedures reveal HRP stained GCs in wholemount views for light microscopical examination. Furthermore, the preservation of the labeled cells and the neuropil is of a quality to allow electron microscopic analysis for synaptic input. This technique can be used in combination with LY backfiling of GCs from the optic nerve and with retinas in which GCs have been prelabeled with rhodamine beads retrogradely transported from the optic tectum as well.

Animals↗