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Synaptic transmission from horizontal cells to cones is impaired by loss of connexin hemichannels.

Klaassen LJ, Sun Z, Steijaert MN, Bolte P, Fahrenfort I, Sjoerdsma T, Klooster J, Claassen Y, Shields CR, Ten Eikelder HM, Janssen-Bienhold U, Zoidl G, McMahon DG, Kamermans M - PLoS Biol. (2011)

Bottom Line: A reduction of feedback was also found when horizontal cells were pharmacologically hyperpolarized but was absent when they were pharmacologically depolarized.A model for feedback, in which the number of connexin hemichannels is reduced to about 40%, fully predicts the specific asymmetric modification of feedback.To our knowledge, this is the first successful genetic interference in the feedback pathway from horizontal cells to cones.

View Article: PubMed Central - PubMed

Affiliation: Research Unit Retinal Signal Processing, The Netherlands Institute for Neuroscience, Amsterdam, The Netherlands.

ABSTRACT
In the vertebrate retina, horizontal cells generate the inhibitory surround of bipolar cells, an essential step in contrast enhancement. For the last decades, the mechanism involved in this inhibitory synaptic pathway has been a major controversy in retinal research. One hypothesis suggests that connexin hemichannels mediate this negative feedback signal; another suggests that feedback is mediated by protons. Mutant zebrafish were generated that lack connexin 55.5 hemichannels in horizontal cells. Whole cell voltage clamp recordings were made from isolated horizontal cells and cones in flat mount retinas. Light-induced feedback from horizontal cells to cones was reduced in mutants. A reduction of feedback was also found when horizontal cells were pharmacologically hyperpolarized but was absent when they were pharmacologically depolarized. Hemichannel currents in isolated horizontal cells showed a similar behavior. The hyperpolarization-induced hemichannel current was strongly reduced in the mutants while the depolarization-induced hemichannel current was not. Intracellular recordings were made from horizontal cells. Consistent with impaired feedback in the mutant, spectral opponent responses in horizontal cells were diminished in these animals. A behavioral assay revealed a lower contrast-sensitivity, illustrating the role of the horizontal cell to cone feedback pathway in contrast enhancement. Model simulations showed that the observed modifications of feedback can be accounted for by an ephaptic mechanism. A model for feedback, in which the number of connexin hemichannels is reduced to about 40%, fully predicts the specific asymmetric modification of feedback. To our knowledge, this is the first successful genetic interference in the feedback pathway from horizontal cells to cones. It provides direct evidence for an unconventional role of connexin hemichannels in the inhibitory synapse between horizontal cells and cones. This is an important step in resolving a long-standing debate about the unusual form of (ephaptic) synaptic transmission between horizontal cells and cones in the vertebrate retina.

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Optokinetic response of mutant zebrafish is reduced.(A) Eye movements of a wild-type zebrafish larva (black, right eye; red, left eye). Timing of the stimulus is indicated in the bottom trace. (B) Optokinetic gain as function of contrast for 13 wild-type (black) and 13 mutant (red) zebrafish. Over the whole contrast range the wild-type performed significantly better than the mutant. (C) In the mutant, reduction in optokinetic gain is stronger for high temporal frequencies (1.0 cycle per second) than for low temporal frequencies (0.25 cycle per second) (p = 0.017). (D) In wild-type the temporal frequency transfer function of cones to bipolar cells is a band-pass filter. The low frequency cutoff is due to negative feedback from horizontal cells. When removing this pathway and inducing an overall gain reduction, the transfer function changes into a low-pass filter. At low frequencies this transformation leads to a smaller loss in gain than for high temporal frequencies.
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pbio-1001107-g009: Optokinetic response of mutant zebrafish is reduced.(A) Eye movements of a wild-type zebrafish larva (black, right eye; red, left eye). Timing of the stimulus is indicated in the bottom trace. (B) Optokinetic gain as function of contrast for 13 wild-type (black) and 13 mutant (red) zebrafish. Over the whole contrast range the wild-type performed significantly better than the mutant. (C) In the mutant, reduction in optokinetic gain is stronger for high temporal frequencies (1.0 cycle per second) than for low temporal frequencies (0.25 cycle per second) (p = 0.017). (D) In wild-type the temporal frequency transfer function of cones to bipolar cells is a band-pass filter. The low frequency cutoff is due to negative feedback from horizontal cells. When removing this pathway and inducing an overall gain reduction, the transfer function changes into a low-pass filter. At low frequencies this transformation leads to a smaller loss in gain than for high temporal frequencies.

Mentions: How would vision be affected by this mutation? We have shown previously that feedback has two components: subtractive and multiplicative [31]. The subtractive component subtracts the global mean activity of the cone output and the multiplicative component amplifies the remaining signal, scaling it such that it fits the bandwidth of the bipolar cells. The result is that the information about objects deviating from the surround is transmitted with high fidelity to the brain. Such a mechanism also generates a form of color-constancy [32],[33]. Without feedback the amount of information transmitted from the cones to the second order neurons would be reduced, thus lowering their contrast sensitivity and color-constancy. To test the first prediction, we measured the optokinetic response (OKR) of 5-d-old zebrafish larvae as function of contrast. OKR responses were generated with an LED-based optokinetic stimulator (Figure S3). This stimulator allows the projection of (sine-wave) patterns of light of various contrasts and colors and rotates these patterns with various velocities. Eye movements are recorded (Figure 9A) and the optokinetic gain is determined by dividing the eye movement velocity by the velocity of the stimulus. Figure 9B shows that for all contrasts the optokinetic gain was significantly reduced in mutant zebrafish compared to wild-type (p<0.01). Since Cx55.5 is exclusively expressed in retinal horizontal cells [7], these experiments show that negative feedback from horizontal cells to cones is important for contrast sensitivity. Finally, in a separate set of experiments, the temporal frequency dependence of the optokinetic gain was determined (Figure 9C). The gain reduction found in the mutants was stronger for high than for low temporal frequencies (p = 0.017). This is consistent with the notion that a negative feedback pathway enhances the synaptic gain [31] and acts as a band-pass filter. Removing negative feedback will reduce the gain and in addition convert the filter into a low pass filter (Figure 9D). The result is that high frequencies will be affected more by the gain reduction than low temporal frequencies.


Synaptic transmission from horizontal cells to cones is impaired by loss of connexin hemichannels.

Klaassen LJ, Sun Z, Steijaert MN, Bolte P, Fahrenfort I, Sjoerdsma T, Klooster J, Claassen Y, Shields CR, Ten Eikelder HM, Janssen-Bienhold U, Zoidl G, McMahon DG, Kamermans M - PLoS Biol. (2011)

Optokinetic response of mutant zebrafish is reduced.(A) Eye movements of a wild-type zebrafish larva (black, right eye; red, left eye). Timing of the stimulus is indicated in the bottom trace. (B) Optokinetic gain as function of contrast for 13 wild-type (black) and 13 mutant (red) zebrafish. Over the whole contrast range the wild-type performed significantly better than the mutant. (C) In the mutant, reduction in optokinetic gain is stronger for high temporal frequencies (1.0 cycle per second) than for low temporal frequencies (0.25 cycle per second) (p = 0.017). (D) In wild-type the temporal frequency transfer function of cones to bipolar cells is a band-pass filter. The low frequency cutoff is due to negative feedback from horizontal cells. When removing this pathway and inducing an overall gain reduction, the transfer function changes into a low-pass filter. At low frequencies this transformation leads to a smaller loss in gain than for high temporal frequencies.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3139627&req=5

pbio-1001107-g009: Optokinetic response of mutant zebrafish is reduced.(A) Eye movements of a wild-type zebrafish larva (black, right eye; red, left eye). Timing of the stimulus is indicated in the bottom trace. (B) Optokinetic gain as function of contrast for 13 wild-type (black) and 13 mutant (red) zebrafish. Over the whole contrast range the wild-type performed significantly better than the mutant. (C) In the mutant, reduction in optokinetic gain is stronger for high temporal frequencies (1.0 cycle per second) than for low temporal frequencies (0.25 cycle per second) (p = 0.017). (D) In wild-type the temporal frequency transfer function of cones to bipolar cells is a band-pass filter. The low frequency cutoff is due to negative feedback from horizontal cells. When removing this pathway and inducing an overall gain reduction, the transfer function changes into a low-pass filter. At low frequencies this transformation leads to a smaller loss in gain than for high temporal frequencies.
Mentions: How would vision be affected by this mutation? We have shown previously that feedback has two components: subtractive and multiplicative [31]. The subtractive component subtracts the global mean activity of the cone output and the multiplicative component amplifies the remaining signal, scaling it such that it fits the bandwidth of the bipolar cells. The result is that the information about objects deviating from the surround is transmitted with high fidelity to the brain. Such a mechanism also generates a form of color-constancy [32],[33]. Without feedback the amount of information transmitted from the cones to the second order neurons would be reduced, thus lowering their contrast sensitivity and color-constancy. To test the first prediction, we measured the optokinetic response (OKR) of 5-d-old zebrafish larvae as function of contrast. OKR responses were generated with an LED-based optokinetic stimulator (Figure S3). This stimulator allows the projection of (sine-wave) patterns of light of various contrasts and colors and rotates these patterns with various velocities. Eye movements are recorded (Figure 9A) and the optokinetic gain is determined by dividing the eye movement velocity by the velocity of the stimulus. Figure 9B shows that for all contrasts the optokinetic gain was significantly reduced in mutant zebrafish compared to wild-type (p<0.01). Since Cx55.5 is exclusively expressed in retinal horizontal cells [7], these experiments show that negative feedback from horizontal cells to cones is important for contrast sensitivity. Finally, in a separate set of experiments, the temporal frequency dependence of the optokinetic gain was determined (Figure 9C). The gain reduction found in the mutants was stronger for high than for low temporal frequencies (p = 0.017). This is consistent with the notion that a negative feedback pathway enhances the synaptic gain [31] and acts as a band-pass filter. Removing negative feedback will reduce the gain and in addition convert the filter into a low pass filter (Figure 9D). The result is that high frequencies will be affected more by the gain reduction than low temporal frequencies.

Bottom Line: A reduction of feedback was also found when horizontal cells were pharmacologically hyperpolarized but was absent when they were pharmacologically depolarized.A model for feedback, in which the number of connexin hemichannels is reduced to about 40%, fully predicts the specific asymmetric modification of feedback.To our knowledge, this is the first successful genetic interference in the feedback pathway from horizontal cells to cones.

View Article: PubMed Central - PubMed

Affiliation: Research Unit Retinal Signal Processing, The Netherlands Institute for Neuroscience, Amsterdam, The Netherlands.

ABSTRACT
In the vertebrate retina, horizontal cells generate the inhibitory surround of bipolar cells, an essential step in contrast enhancement. For the last decades, the mechanism involved in this inhibitory synaptic pathway has been a major controversy in retinal research. One hypothesis suggests that connexin hemichannels mediate this negative feedback signal; another suggests that feedback is mediated by protons. Mutant zebrafish were generated that lack connexin 55.5 hemichannels in horizontal cells. Whole cell voltage clamp recordings were made from isolated horizontal cells and cones in flat mount retinas. Light-induced feedback from horizontal cells to cones was reduced in mutants. A reduction of feedback was also found when horizontal cells were pharmacologically hyperpolarized but was absent when they were pharmacologically depolarized. Hemichannel currents in isolated horizontal cells showed a similar behavior. The hyperpolarization-induced hemichannel current was strongly reduced in the mutants while the depolarization-induced hemichannel current was not. Intracellular recordings were made from horizontal cells. Consistent with impaired feedback in the mutant, spectral opponent responses in horizontal cells were diminished in these animals. A behavioral assay revealed a lower contrast-sensitivity, illustrating the role of the horizontal cell to cone feedback pathway in contrast enhancement. Model simulations showed that the observed modifications of feedback can be accounted for by an ephaptic mechanism. A model for feedback, in which the number of connexin hemichannels is reduced to about 40%, fully predicts the specific asymmetric modification of feedback. To our knowledge, this is the first successful genetic interference in the feedback pathway from horizontal cells to cones. It provides direct evidence for an unconventional role of connexin hemichannels in the inhibitory synapse between horizontal cells and cones. This is an important step in resolving a long-standing debate about the unusual form of (ephaptic) synaptic transmission between horizontal cells and cones in the vertebrate retina.

Show MeSH
Related in: MedlinePlus