Synaptotagmin I Regulates Patterned Spontaneous Activity in the Developing Rat Retina via Calcium Binding to the C2AB Domains
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{"title"=>"Synaptotagmin I Regulates Patterned Spontaneous Activity in the Developing Rat Retina via Calcium Binding to the C2AB Domains", "type"=>"journal", "authors"=>[{"first_name"=>"Chung Wei", "last_name"=>"Chiang", "scopus_author_id"=>"55414334600"}, {"first_name"=>"Yu Chieh", "last_name"=>"Chen", "scopus_author_id"=>"36175438200"}, {"first_name"=>"Juu Chin", "last_name"=>"Lu", "scopus_author_id"=>"7601564535"}, {"first_name"=>"Yu Tien", "last_name"=>"Hsiao", "scopus_author_id"=>"55413235300"}, {"first_name"=>"Che Wei", "last_name"=>"Chang", "scopus_author_id"=>"37048378900"}, {"first_name"=>"Pin Chien", "last_name"=>"Huang", "scopus_author_id"=>"55413301100"}, {"first_name"=>"Yu Tzu", "last_name"=>"Chang", "scopus_author_id"=>"55816134400"}, {"first_name"=>"Payne Y.", "last_name"=>"Chang", "scopus_author_id"=>"7403573665"}, {"first_name"=>"Chih Tien", "last_name"=>"Wang", "scopus_author_id"=>"8072034100"}], "year"=>2012, "source"=>"PLoS ONE", "identifiers"=>{"pui"=>"365872501", "pmid"=>"23091625", "isbn"=>"1932-6203 (Electronic)\\r1932-6203 (Linking)", "issn"=>"19326203", "doi"=>"10.1371/journal.pone.0047465", "scopus"=>"2-s2.0-84867617097", "sgr"=>"84867617097"}, "id"=>"46f27cde-b4f2-3d3e-a8f3-3c9233b2138a", "abstract"=>"BACKGROUND: In neonatal binocular animals, the developing retina displays patterned spontaneous activity termed retinal waves, which are initiated by a single class of interneurons (starburst amacrine cells, SACs) that release neurotransmitters. Although SACs are shown to regulate wave dynamics, little is known regarding how altering the proteins involved in neurotransmitter release may affect wave dynamics. Synaptotagmin (Syt) family harbors two Ca(2+)-binding domains (C2A and C2B) which serve as Ca(2+) sensors in neurotransmitter release. However, it remains unclear whether SACs express any specific Syt isoform mediating retinal waves. Moreover, it is unknown how Ca(2+) binding to C2A and C2B of Syt affects wave dynamics. Here, we investigated the expression of Syt I in the neonatal rat retina and examined the roles of C2A and C2B in regulating wave dynamics. METHODOLOGY/PRINCIPAL FINDINGS: Immunostaining and confocal microscopy showed that Syt I was expressed in neonatal rat SACs and cholinergic synapses, consistent with its potential role as a Ca(2+) sensor mediating retinal waves. By combining a horizontal electroporation strategy with the SAC-specific promoter, we specifically expressed Syt I mutants with weakened Ca(2+)-binding ability in C2A or C2B in SACs. Subsequent live Ca(2+) imaging was used to monitor the effects of these molecular perturbations on wave-associated spontaneous Ca(2+) transients. We found that targeted expression of Syt I C2A or C2B mutants in SACs significantly reduced the frequency, duration, and amplitude of wave-associated Ca(2+) transients, suggesting that both C2 domains regulate wave temporal properties. In contrast, these C2 mutants had relatively minor effects on pairwise correlations over distance for wave-associated Ca(2+) transients. CONCLUSIONS/SIGNIFICANCE: Through Ca(2+) binding to C2A or C2B, the Ca(2+) sensor Syt I in SACs may regulate patterned spontaneous activity to shape network activity during development. Hence, modulating the releasing machinery in presynaptic neurons (SACs) alters wave dynamics.", "link"=>"http://www.mendeley.com/research/synaptotagmin-i-regulates-patterned-spontaneous-activity-developing-rat-retina-via-calcium-binding-c", "reader_count"=>3, "reader_count_by_academic_status"=>{"Researcher"=>1, "Student > Ph. D. Student"=>1, "Student > Master"=>1}, "reader_count_by_user_role"=>{"Researcher"=>1, "Student > Ph. D. Student"=>1, "Student > Master"=>1}, "reader_count_by_subject_area"=>{"Agricultural and Biological Sciences"=>1, "Medicine and Dentistry"=>1, "Neuroscience"=>1}, "reader_count_by_subdiscipline"=>{"Medicine and Dentistry"=>{"Medicine and Dentistry"=>1}, "Neuroscience"=>{"Neuroscience"=>1}, "Agricultural and Biological Sciences"=>{"Agricultural and Biological Sciences"=>1}}, "group_count"=>0}

Scopus | Further Information

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Figshare

  • {"files"=>["https://ndownloader.figshare.com/files/557870"], "description"=>"<p>Retinal waves were measured from cultured whole-mount retinas expressing Syt I or its mutants (Syt I-C2A* and Syt I-C2B*) with the mGluR2 promoter. Control was transfected with pmGluR2-IRES2EGFP. Wave correlograms were constructed from 49 cell pairs in one imaged region and fitted to a Gaussian equation to yield the Gaussian mean and standard deviation (SD). No significant differences were found in Gaussian mean (0.00±0.00 sec) among all groups. For Gaussian SD, <i>p</i> = 0.6225 (One-Way ANOVA with Student-Newman <i>post-hoc</i> test).</p>", "links"=>[], "tags"=>["correlograms", "syt", "c2ab"], "article_id"=>228357, "categories"=>["Physiology", "Neuroscience", "Developmental Biology"], "users"=>["Chung-Wei Chiang", "Yu-Chieh Chen", "Juu-Chin Lu", "Yu-Tien Hsiao", "Che-Wei Chang", "Pin-Chien Huang", "Yu-Tzu Chang", "Payne Y. Chang", "Chih-Tien Wang"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0047465.t002", "stats"=>{"downloads"=>0, "page_views"=>4, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Comparison_of_wave_correlograms_for_Syt_I_and_its_C2AB_mutants_/228357", "title"=>"Comparison of wave correlograms for Syt I and its C2AB mutants.", "pos_in_sequence"=>0, "defined_type"=>3, "published_date"=>"2012-10-16 02:19:17"}
  • {"files"=>["https://ndownloader.figshare.com/files/557689"], "description"=>"<p>A. <i>Top,</i> Definitions of Ca<sup>2+</sup> transient duration and amplitude. The peaks of Ca<sup>2+</sup> transients were automatically picked by Igor Pro according to the criteria in <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0047465#s4\" target=\"_blank\">Materials and Methods</a>. The RMS noise (Noise) was measured from the trace between 30 sec prior to the peak and 50 sec following the peak. The starting point (x<sub>0</sub>, y<sub>0</sub>) (indicated by solid arrows) was defined by the latest time point where the first derivative was zero (i.e. dy/dx  = 0, where y was the fluorescence changes in % ΔF/F and x was the recording time in sec) before the peak. To define the end point (x′, y′) (indicated by open arrows), a line was drawn to connect the time points where the trace of Ca<sup>2+</sup> transients returned to within the RMS noise of the baseline, with the minimal fluctuation of fluorescence between the starting and end points (i.e. y′−y<sub>0</sub> = minimum). <i>Bottom,</i> The Ca<sup>2+</sup> transient duration was the interval between the starting and end points. The Ca<sup>2+</sup> transient amplitude was the fluorescence change from the baseline to peak. B. Representative individual Ca<sup>2+</sup> transients from retinas transfected with Control (black), Syt I (blue), Syt I-C2A* (green), or Syt I-C2B* (red). C. Summary of Ca<sup>2+</sup> transient duration for each group. Data were from 22–39 transfected retinas and 7–15 pups (about 50% data in each group from the retinas transfected on P0 with DIV 3–4). (*<i>p</i><0.05; **<i>p</i><0.01; ***<i>p</i><0.001; Kruskal-Wallis method followed by <i>post</i>-<i>hoc</i> Dunn test.) D. Distributions of cumulative probability for Ca<sup>2+</sup> transient duration from individual cells. Data were from 860–1106 cells out of the same data sets in C. E. Summary of Ca<sup>2+</sup> transient amplitude after transfection. Data sets were the same as in C. (**<i>p</i><0.01; Kruskal-Wallis method followed by <i>post</i>-<i>hoc</i> Dunn test.) F. Distributions of cumulative probability for Ca<sup>2+</sup> transient amplitude from individual cells. Data sets were the same as in D.</p>", "links"=>[], "tags"=>["transient", "reduced", "weakened", "binding", "c2ab", "domains", "syt"], "article_id"=>228168, "categories"=>["Physiology", "Neuroscience", "Developmental Biology"], "users"=>["Chung-Wei Chiang", "Yu-Chieh Chen", "Juu-Chin Lu", "Yu-Tien Hsiao", "Che-Wei Chang", "Pin-Chien Huang", "Yu-Tzu Chang", "Payne Y. Chang", "Chih-Tien Wang"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0047465.g005", "stats"=>{"downloads"=>0, "page_views"=>3, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Ca_2_transient_size_is_reduced_by_weakened_Ca_2_binding_to_the_C2AB_domains_of_Syt_I_/228168", "title"=>"Ca<sup>2+</sup> transient size is reduced by weakened Ca<sup>2+</sup> binding to the C2AB domains of Syt I.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-10-16 02:16:08"}
  • {"files"=>["https://ndownloader.figshare.com/files/557432"], "description"=>"<p>Whole-mount retinas from P0–P2 rats were double-labeled with HA (green) and ChAT (red) after 72 hr of transfection. Ai–Ci. Whole-mount retinas transfected with pCMV-HA-Syt I-IRES2-EGFP. Ai. Labeled with HA. Bi. Labeled with ChAT. Ci. Merged images of Ai and Bi. Scale bar, 50 µm. EGFP, enhanced green fluorescent protein. Aii–Cii. High magnification view of whole-mount retinas transfected with pCMV-HA-Syt I-IRES2-EGFP. Aii. Labeled with HA. Bii. Labeled with ChAT. Cii. Merged images of Aii and Bii. Scale bar, 25 µm. Di–Fi. Whole-mount retinas transfected with pmGluR2-HA-Syt I-IRES2-EGFP. Di. Labeled with HA. Ei. Labeled with ChAT. Fi. Merged images of Di and Ei. Scale bar, 50 µm. Dii-Fii. High magnification view of whole-mount retinas transfected with pmGluR2-HA-Syt I-IRES2-EGFP. Dii. Labeled with HA. Eii. Labeled with ChAT. Fii. Merged images of Dii and Eii. Scale bar, 25 µm. G–H. Retinal cross-sections transfected with either pCMV-HA-Syt I-IRES2-EGFP (G), or pmGluR2-HA-Syt I-IRES2-EGFP (H). The SACs and IPL were stained for ChAT (red). The transfected cells were stained for HA/GFP (green). The cell nuclei were stained with DAPI (blue). <i>Right,</i> Merged images (yellow). The transfected SACs were indicated by arrows. <i>Insets</i> showed the higher magnification of merged images. Scale bars, 25 µm.</p>", "links"=>[], "tags"=>["mglur2", "promoter", "drives", "selectively"], "article_id"=>227914, "categories"=>["Physiology", "Neuroscience", "Developmental Biology"], "users"=>["Chung-Wei Chiang", "Yu-Chieh Chen", "Juu-Chin Lu", "Yu-Tien Hsiao", "Che-Wei Chang", "Pin-Chien Huang", "Yu-Tzu Chang", "Payne Y. Chang", "Chih-Tien Wang"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0047465.g003", "stats"=>{"downloads"=>1, "page_views"=>5, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_The_mGluR2_promoter_drives_gene_expression_selectively_in_SACs_/227914", "title"=>"The mGluR2 promoter drives gene expression selectively in SACs.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-10-16 02:11:54"}
  • {"files"=>["https://ndownloader.figshare.com/files/557147"], "description"=>"<p>Ai–Aiii. Merged images of Syt I (green) and ChAT (red) staining in retinal cross-sections from P0–P2 rats. The colocalization of Syt I and ChAT labeling was found in the IPL (yellow). Scale bars, 25 µm. ChAT, choline acetyltransferase. NBL, neuroblast layer; IPL, inner plexiform layer; GCL, ganglion cell layer. Bi–Biii. Immunofluorescence labeling of Syt I (green) in the IPL of retinal cross-sections from P0–P2 rats. Ci–Ciii. Immunofluorescence staining of ChAT (red) labeling SACs and the IPL in retinal cross-sections from P0–P2 rats. Di–Diii. Merged images of Syt I and ChAT staining in the same retinal cross-sections. The Syt I and ChAT labeling was colocalized to the SAC somata (yellow) as indicated by arrows. Scale bars for B-D, 50 µm. Ei–Eiii. The high magnification for merged images of Syt I and ChAT staining in the boxes of Di–Diii. Scale bars for E, 5 µm.</p>", "links"=>[], "tags"=>["neonatal", "sacs"], "article_id"=>227631, "categories"=>["Physiology", "Neuroscience", "Developmental Biology"], "users"=>["Chung-Wei Chiang", "Yu-Chieh Chen", "Juu-Chin Lu", "Yu-Tien Hsiao", "Che-Wei Chang", "Pin-Chien Huang", "Yu-Tzu Chang", "Payne Y. Chang", "Chih-Tien Wang"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0047465.g001", "stats"=>{"downloads"=>1, "page_views"=>4, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Syt_I_is_strongly_expressed_in_neonatal_SACs_and_IPL_/227631", "title"=>"Syt I is strongly expressed in neonatal SACs and IPL.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-10-16 02:07:11"}
  • {"files"=>["https://ndownloader.figshare.com/files/557911"], "description"=>"<p>Retinal waves were measured from whole-mount retinal explant cultures after transfection with the DNA plasmid (pmGluR2-IRES2EGFP). Intact control was P0–P2 retinas in the same culture condition (DIV 3–4) without transfection. No significant differences were found among all groups. For wave frequency, <i>p</i> = 0.26 (Kruskal-Wallis method with Dunn <i>post-hoc</i> test); for wave duration, <i>p</i> = 0.58 (One-way ANOVA with Student-Newman <i>post-hoc</i> test); for wave amplitude, <i>p</i> = 0.87 (One-way ANOVA with Student-Newman <i>post-hoc</i> test).</p>", "links"=>[], "tags"=>["characteristics"], "article_id"=>228404, "categories"=>["Physiology", "Neuroscience", "Developmental Biology"], "users"=>["Chung-Wei Chiang", "Yu-Chieh Chen", "Juu-Chin Lu", "Yu-Tien Hsiao", "Che-Wei Chang", "Pin-Chien Huang", "Yu-Tzu Chang", "Payne Y. Chang", "Chih-Tien Wang"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0047465.t001", "stats"=>{"downloads"=>1, "page_views"=>5, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Comparison_of_wave_characteristics_following_transfection_/228404", "title"=>"Comparison of wave characteristics following transfection.", "pos_in_sequence"=>0, "defined_type"=>3, "published_date"=>"2012-10-16 02:20:04"}
  • {"files"=>["https://ndownloader.figshare.com/files/557580"], "description"=>"<p>A. <i>Left,</i> The expression pattern of pmGluR2-IRES2EGFP in transfected whole-amount retinas. <i>Right,</i> The RGC layer labeled with the Ca<sup>2+</sup> indicator fura-2 to measure the wave-associated Ca<sup>2+</sup> transients after transfection. Scale bar for both panels, 25 µm. B. Spontaneous, correlated Ca<sup>2+</sup> transients were monitored from randomly selected cells in the RGC layer. The example traces of fluorescent changes over time showed the Ca<sup>2+</sup> transients in the nearby cells from one imaged region. Retinas were transfected with pmGluR2-IRES2EGFP (Control), pmGluR2-IRES2EGFP-Syt I (Syt I), pmGluR2-IRES2EGFP-Syt I-D230S (Syt I-C2A*), or pmGluR2-IRES2EGFP-Syt I-D363N (Syt I-C2B*). C. Summary of Ca<sup>2+</sup> transient frequency after transfection. Data were from 23–39 transfected retinas and 7–15 pups (about 50% data in each group from the retinas transfected on P0 with DIV 3–4). (**<i>p</i><0.01; ***<i>p</i><0.001; One-Way ANOVA with <i>post</i>-<i>hoc</i> Student-Newman-Keuls test.) D. Distributions of cumulative probability for Ca<sup>2+</sup> transient frequency from individual cells. Data were from 1196–1489 cells out of the same data sets in C. E. Summary of fraction of cells with Ca<sup>2+</sup> transients in one imaged region after transfection. Data were from 32–58 transfected regions out of the same data sets in C. (**<i>p</i><0.01; ***<i>p</i><0.001; Kruskal-Wallis method followed by <i>post</i>-<i>hoc</i> Dunn test.).</p>", "links"=>[], "tags"=>["transient", "reduced", "weakened", "binding", "c2ab", "domains", "syt"], "article_id"=>228053, "categories"=>["Physiology", "Neuroscience", "Developmental Biology"], "users"=>["Chung-Wei Chiang", "Yu-Chieh Chen", "Juu-Chin Lu", "Yu-Tien Hsiao", "Che-Wei Chang", "Pin-Chien Huang", "Yu-Tzu Chang", "Payne Y. Chang", "Chih-Tien Wang"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0047465.g004", "stats"=>{"downloads"=>1, "page_views"=>2, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Ca_2_transient_frequency_is_reduced_by_weakened_Ca_2_binding_to_the_C2AB_domains_of_Syt_I_/228053", "title"=>"Ca<sup>2+</sup> transient frequency is reduced by weakened Ca<sup>2+</sup> binding to the C2AB domains of Syt I.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-10-16 02:14:13"}
  • {"files"=>["https://ndownloader.figshare.com/files/557313"], "description"=>"<p>A–B. Preparation of platinum (Pt) electrodes. Ai. The arrangement of the six slides for the (+) electrode base. The colors of slides represent the slide arrangement in different layers. The numbers in the circles indicate the corresponding sequence to arrange slides. Aii. The Pt foil (15×15 mm with one extra 5×5 mm-overhanging square) was aligned to one side of the 7<sup>th</sup> slide with the overhanging square left out. Epoxy glue was applied to attach the Pt foil onto the slide and connect all the slides together. Aiii. The wire was soldered to the edge of the overhanging Pt square. Aiv. The last (8<sup>th</sup>) slide was glued onto the (+) electrode base. Bi–ii. The arrangement of the (−) electrode. A pen tube (green) with a diameter of 10 mm was used to attach the same-sized Pt foil by epoxy glue. The electric wire was inserted through the pen tube and soldered onto the Pt foil. C. The setup for electroporation in a horizontal configuration. The retinal explant was placed in a well [with dimensions 12 (length) ×12 (width) ×3 (height) mm] on the (+) electrode. The (−) electrode made contact with solution above the well and covered the retinal explant. The distance between the (+) and (−) electrodes was adjusted by a micromanipulator that held the (−) electrode. D. The P1 rat retinas were transfected with pCMV-HA (vector) or pCMV-HA-Syt I (HA-Syt I) with this electroporation device. The retinal explants were incubated for 72 hr to allow gene expression. Cellular proteins were solubilized and subjected to SDS-PAGE and Western blot analysis with antibodies indicated on the right (HA or α-tubulin). Only the retinas transfected with HA-Syt I displayed the HA signal with a size of ∼65 kD, which corresponded to the molecular weight of Syt I. Data shown were representative blots from 3 different experiments.</p>", "links"=>[], "tags"=>["whole-mount", "retinas", "homemade", "electroporation"], "article_id"=>227791, "categories"=>["Physiology", "Neuroscience", "Developmental Biology"], "users"=>["Chung-Wei Chiang", "Yu-Chieh Chen", "Juu-Chin Lu", "Yu-Tien Hsiao", "Che-Wei Chang", "Pin-Chien Huang", "Yu-Tzu Chang", "Payne Y. Chang", "Chih-Tien Wang"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0047465.g002", "stats"=>{"downloads"=>1, "page_views"=>2, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Gene_transfer_into_whole_mount_retinas_by_the_homemade_electroporation_device_/227791", "title"=>"Gene transfer into whole-mount retinas by the homemade electroporation device.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-10-16 02:09:51"}
  • {"files"=>["https://ndownloader.figshare.com/files/557788"], "description"=>"<p>A. Representative pairwise cross-correlograms of 49 cells in one imaged region from the transfected retinas. Each event was a Ca<sup>2+</sup> transient in an individual cell. Delay times were the time differences between Ca<sup>2+</sup> transient peaks in cell pairs. Note that all correlograms for control, Syt I, Syt I-C2A* and Syt I-C2B* were very similar with sharp peaks at t = 0 sec (see <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0047465#pone-0047465-t002\" target=\"_blank\">Table 2</a> for statistics). The oscillatory behavior of the correlograms beyond | t | > ∼7.5 sec might reflect the bursting behavior of the distant cells. Bin widths for all correlograms were 1 sec. B. Pairwise C.I. values as a function of intercellular distance for control, Syt I, Syt I-C2A* and Syt I-C2B*. Data points were averages of medians within the 50 µm bins from 17–23 transfected retinas (about 50% data in each group from the retinas transfected on P0 with DIV 3–4). The error bars were the standard errors of the C.I. values in each bin.</p>", "links"=>[], "tags"=>["altered", "weakened", "binding", "c2ab", "syt"], "article_id"=>228266, "categories"=>["Physiology", "Neuroscience", "Developmental Biology"], "users"=>["Chung-Wei Chiang", "Yu-Chieh Chen", "Juu-Chin Lu", "Yu-Tien Hsiao", "Che-Wei Chang", "Pin-Chien Huang", "Yu-Tzu Chang", "Payne Y. Chang", "Chih-Tien Wang"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0047465.g006", "stats"=>{"downloads"=>1, "page_views"=>9, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Pairwise_correlation_is_not_altered_by_weakened_Ca_2_binding_to_the_C2AB_domain_of_Syt_I_/228266", "title"=>"Pairwise correlation is not altered by weakened Ca<sup>2+</sup> binding to the C2AB domain of Syt I.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-10-16 02:17:46"}
  • {"files"=>["https://ndownloader.figshare.com/files/296790"], "description"=>"<div><h3>Background</h3><p>In neonatal binocular animals, the developing retina displays patterned spontaneous activity termed retinal waves, which are initiated by a single class of interneurons (starburst amacrine cells, SACs) that release neurotransmitters. Although SACs are shown to regulate wave dynamics, little is known regarding how altering the proteins involved in neurotransmitter release may affect wave dynamics. Synaptotagmin (Syt) family harbors two Ca<sup>2+</sup>-binding domains (C2A and C2B) which serve as Ca<sup>2+</sup> sensors in neurotransmitter release. However, it remains unclear whether SACs express any specific Syt isoform mediating retinal waves. Moreover, it is unknown how Ca<sup>2+</sup> binding to C2A and C2B of Syt affects wave dynamics. Here, we investigated the expression of Syt I in the neonatal rat retina and examined the roles of C2A and C2B in regulating wave dynamics.</p> <h3>Methodology/Principal Findings</h3><p>Immunostaining and confocal microscopy showed that Syt I was expressed in neonatal rat SACs and cholinergic synapses, consistent with its potential role as a Ca<sup>2+</sup> sensor mediating retinal waves. By combining a horizontal electroporation strategy with the SAC-specific promoter, we specifically expressed Syt I mutants with weakened Ca<sup>2+</sup>-binding ability in C2A or C2B in SACs. Subsequent live Ca<sup>2+</sup> imaging was used to monitor the effects of these molecular perturbations on wave-associated spontaneous Ca<sup>2+</sup> transients. We found that targeted expression of Syt I C2A or C2B mutants in SACs significantly reduced the frequency, duration, and amplitude of wave-associated Ca<sup>2+</sup> transients, suggesting that both C2 domains regulate wave temporal properties. In contrast, these C2 mutants had relatively minor effects on pairwise correlations over distance for wave-associated Ca<sup>2+</sup> transients.</p> <h3>Conclusions/Significance</h3><p>Through Ca<sup>2+</sup> binding to C2A or C2B, the Ca<sup>2+</sup> sensor Syt I in SACs may regulate patterned spontaneous activity to shape network activity during development. Hence, modulating the releasing machinery in presynaptic neurons (SACs) alters wave dynamics.</p> </div>", "links"=>[], "tags"=>["synaptotagmin", "regulates", "patterned", "spontaneous", "retina", "calcium", "binding", "c2ab", "domains"], "article_id"=>118479, "categories"=>["Physiology", "Neuroscience", "Developmental Biology"], "users"=>["Chung-Wei Chiang", "Yu-Chieh Chen", "Juu-Chin Lu", "Yu-Tien Hsiao", "Che-Wei Chang", "Pin-Chien Huang", "Yu-Tzu Chang", "Payne Y. Chang", "Chih-Tien Wang"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0047465", "stats"=>{"downloads"=>1, "page_views"=>21, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/Synaptotagmin_I_Regulates_Patterned_Spontaneous_Activity_in_the_Developing_Rat_Retina_via_Calcium_Binding_to_the_C2AB_Domains/118479", "title"=>"Synaptotagmin I Regulates Patterned Spontaneous Activity in the Developing Rat Retina via Calcium Binding to the C2AB Domains", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-10-16 02:21:19"}

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