GluA3 subunits are required for appropriate assembly of AMPAR GluA2 and GluA4 subunits on cochlear afferent synapses and for presynaptic ribbon modiolar–pillar morphology
Cochlear sound encoding depends on α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors (AMPARs), but reliance on specific pore-forming subunits is unknown. With 5-week-old male C57BL/6J Gria3-knockout mice (i.e., subunit GluA3KO) we determined cochlear function, synapse ultrastructure, a...
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| Format: | Article |
| Language: | English |
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eLife Sciences Publications Ltd
2023-01-01
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| Series: | eLife |
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| Online Access: | https://elifesciences.org/articles/80950 |
| _version_ | 1828047332283777024 |
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| author | Mark A Rutherford Atri Bhattacharyya Maolei Xiao Hou-Ming Cai Indra Pal Maria Eulalia Rubio |
| author_facet | Mark A Rutherford Atri Bhattacharyya Maolei Xiao Hou-Ming Cai Indra Pal Maria Eulalia Rubio |
| author_sort | Mark A Rutherford |
| collection | DOAJ |
| description | Cochlear sound encoding depends on α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors (AMPARs), but reliance on specific pore-forming subunits is unknown. With 5-week-old male C57BL/6J Gria3-knockout mice (i.e., subunit GluA3KO) we determined cochlear function, synapse ultrastructure, and AMPAR molecular anatomy at ribbon synapses between inner hair cells (IHCs) and spiral ganglion neurons. GluA3KO and wild-type (GluA3WT) mice reared in ambient sound pressure level (SPL) of 55–75 dB had similar auditory brainstem response (ABR) thresholds, wave-1 amplitudes, and latencies. Postsynaptic densities (PSDs), presynaptic ribbons, and synaptic vesicle sizes were all larger on the modiolar side of the IHCs from GluA3WT, but not GluA3KO, demonstrating GluA3 is required for modiolar–pillar synapse differentiation. Presynaptic ribbons juxtaposed with postsynaptic GluA2/4 subunits were similar in quantity, however, lone ribbons were more frequent in GluA3KO and GluA2-lacking synapses were observed only in GluA3KO. GluA2 and GluA4 immunofluorescence volumes were smaller on the pillar side than the modiolar side in GluA3KO, despite increased pillar-side PSD size. Overall, the fluorescent puncta volumes of GluA2 and GluA4 were smaller in GluA3KO than GluA3WT. However, GluA3KO contained less GluA2 and greater GluA4 immunofluorescence intensity relative to GluA3WT (threefold greater mean GluA4:GluA2 ratio). Thus, GluA3 is essential in development, as germline disruption of Gria3 caused anatomical synapse pathology before cochlear output became symptomatic by ABR. We propose the hearing loss in older male GluA3KO mice results from progressive synaptopathy evident in 5-week-old mice as decreased abundance of GluA2 subunits and an increase in GluA2-lacking, GluA4-monomeric Ca2+-permeable AMPARs. |
| first_indexed | 2024-04-10T18:39:44Z |
| format | Article |
| id | doaj.art-91d0335f112a40cfaafc82ab2015e36e |
| institution | Directory Open Access Journal |
| issn | 2050-084X |
| language | English |
| last_indexed | 2024-04-10T18:39:44Z |
| publishDate | 2023-01-01 |
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| spelling | doaj.art-91d0335f112a40cfaafc82ab2015e36e2023-02-01T18:01:17ZengeLife Sciences Publications LtdeLife2050-084X2023-01-011210.7554/eLife.80950GluA3 subunits are required for appropriate assembly of AMPAR GluA2 and GluA4 subunits on cochlear afferent synapses and for presynaptic ribbon modiolar–pillar morphologyMark A Rutherford0https://orcid.org/0000-0002-2627-6254Atri Bhattacharyya1Maolei Xiao2Hou-Ming Cai3Indra Pal4Maria Eulalia Rubio5https://orcid.org/0000-0003-3536-6013Department of Otolaryngology, Washington University School of Medicine, St Louis, United StatesDepartment of Otolaryngology, Washington University School of Medicine, St Louis, United StatesDepartment of Otolaryngology, Washington University School of Medicine, St Louis, United StatesDepartment of Neurobiology, University of Pittsburgh School of Medicine, Pittsburgh, United StatesDepartment of Neurobiology, University of Pittsburgh School of Medicine, Pittsburgh, United StatesDepartment of Neurobiology, University of Pittsburgh School of Medicine, Pittsburgh, United States; Department of Otolaryngology, University of Pittsburgh School of Medicine, Pittsburgh, United StatesCochlear sound encoding depends on α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors (AMPARs), but reliance on specific pore-forming subunits is unknown. With 5-week-old male C57BL/6J Gria3-knockout mice (i.e., subunit GluA3KO) we determined cochlear function, synapse ultrastructure, and AMPAR molecular anatomy at ribbon synapses between inner hair cells (IHCs) and spiral ganglion neurons. GluA3KO and wild-type (GluA3WT) mice reared in ambient sound pressure level (SPL) of 55–75 dB had similar auditory brainstem response (ABR) thresholds, wave-1 amplitudes, and latencies. Postsynaptic densities (PSDs), presynaptic ribbons, and synaptic vesicle sizes were all larger on the modiolar side of the IHCs from GluA3WT, but not GluA3KO, demonstrating GluA3 is required for modiolar–pillar synapse differentiation. Presynaptic ribbons juxtaposed with postsynaptic GluA2/4 subunits were similar in quantity, however, lone ribbons were more frequent in GluA3KO and GluA2-lacking synapses were observed only in GluA3KO. GluA2 and GluA4 immunofluorescence volumes were smaller on the pillar side than the modiolar side in GluA3KO, despite increased pillar-side PSD size. Overall, the fluorescent puncta volumes of GluA2 and GluA4 were smaller in GluA3KO than GluA3WT. However, GluA3KO contained less GluA2 and greater GluA4 immunofluorescence intensity relative to GluA3WT (threefold greater mean GluA4:GluA2 ratio). Thus, GluA3 is essential in development, as germline disruption of Gria3 caused anatomical synapse pathology before cochlear output became symptomatic by ABR. We propose the hearing loss in older male GluA3KO mice results from progressive synaptopathy evident in 5-week-old mice as decreased abundance of GluA2 subunits and an increase in GluA2-lacking, GluA4-monomeric Ca2+-permeable AMPARs.https://elifesciences.org/articles/80950glutamate receptorsinner hair cellsconfocal imaging3D reconstructionsqRT-PCRimmunolabeling |
| spellingShingle | Mark A Rutherford Atri Bhattacharyya Maolei Xiao Hou-Ming Cai Indra Pal Maria Eulalia Rubio GluA3 subunits are required for appropriate assembly of AMPAR GluA2 and GluA4 subunits on cochlear afferent synapses and for presynaptic ribbon modiolar–pillar morphology eLife glutamate receptors inner hair cells confocal imaging 3D reconstructions qRT-PCR immunolabeling |
| title | GluA3 subunits are required for appropriate assembly of AMPAR GluA2 and GluA4 subunits on cochlear afferent synapses and for presynaptic ribbon modiolar–pillar morphology |
| title_full | GluA3 subunits are required for appropriate assembly of AMPAR GluA2 and GluA4 subunits on cochlear afferent synapses and for presynaptic ribbon modiolar–pillar morphology |
| title_fullStr | GluA3 subunits are required for appropriate assembly of AMPAR GluA2 and GluA4 subunits on cochlear afferent synapses and for presynaptic ribbon modiolar–pillar morphology |
| title_full_unstemmed | GluA3 subunits are required for appropriate assembly of AMPAR GluA2 and GluA4 subunits on cochlear afferent synapses and for presynaptic ribbon modiolar–pillar morphology |
| title_short | GluA3 subunits are required for appropriate assembly of AMPAR GluA2 and GluA4 subunits on cochlear afferent synapses and for presynaptic ribbon modiolar–pillar morphology |
| title_sort | glua3 subunits are required for appropriate assembly of ampar glua2 and glua4 subunits on cochlear afferent synapses and for presynaptic ribbon modiolar pillar morphology |
| topic | glutamate receptors inner hair cells confocal imaging 3D reconstructions qRT-PCR immunolabeling |
| url | https://elifesciences.org/articles/80950 |
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