In dopaminergic (DA) substantia nigra (SN) neurons Cav2.3 R-type Ca2+-currents contribute to somatodendritic Ca2+-oscillations. This activity may contribute to the selective degeneration of these neurons in Parkinson's disease (PD) since Cav2.3-knockout is neuroprotective in a PD mouse model. Here we show that in tsA-201-cells the membrane-anchored β2-splice variants β2a and β2e are required to stabilize Cav2.3 gating properties allowing sustained Cav2.3 availability during simulated pacemaking and enhanced Ca2+-currents during bursts. We confirmed the expression of β2a- and β2e-subunit transcripts in the mouse SN and in identified SN DA neurons. Patch-clamp recordings of mouse DA midbrain neurons in culture and SN DA neurons in brain slices revealed SNX-482-sensitive R-type Ca2+-currents with voltage-dependent gating properties that suggest modulation by β2a- and/or β2e-subunits. Thus, β-subunit alternative splicing may prevent a fraction of Cav2.3 channels from inactivation in continuously active, highly vulnerable SN DA neurons, thereby also supporting Ca2+ signals contributing to the (patho)physiological role of Cav2.3 channels in PD.
All data generated or analyzed during this study are included in the manuscript and supporting files. Raw data have been provided for mean population data shown in Figures and Tables.
RNASeq of DA neurons from SNpc and VTA. Dataset posted on 04.03.2015, 16:01 by William Shindoi.org/10.6084/m9.figshare.926519.v1.
RNA Sequencing Identifies Robust Markers of Vulnerable and Resistant Human Midbrain Dopamine Neurons and Their Expression in Parkinson's DiseaseNCBI Gene Expression Omnibus, GSE114918.
- Jörg Striessnig
- Nadine Jasmin Ortner
- Emilio Carbone
- Emilio Carbone
- Nadine Jasmin Ortner
- Birgit Liss
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Animal experimentation: All animal experiments and procedures were performed in strict accordance with the European Community's Council Directive 2010/63/UE and approved by the Italian Ministry of Health and the Local Organism responsible for animal welfare at the University of Torino (authorization DGSAF 0011710-P-26/07/2017) and the local authorities at the University of Ulm (Regierungspräsidium Tübingen, Ref: 35/9185.81-3; Reg. Nr. o.147) and University of Cologne (LANUV NRW, Recklinghausen, Germany (84-02.05.20.12.254).
- Henry M Colecraft, Columbia University, United States
- Preprint posted: February 10, 2021 (view preprint)
- Received: February 11, 2021
- Accepted: July 4, 2022
- Accepted Manuscript published: July 6, 2022 (version 1)
- Accepted Manuscript updated: July 8, 2022 (version 2)
- Version of Record published: July 22, 2022 (version 3)
- Version of Record updated: July 27, 2022 (version 4)
© 2022, Siller et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Deciphering patterns of connectivity between neurons in the brain is a critical step toward understanding brain function. Imaging-based neuroanatomical tracing identifies area-to-area or sparse neuron-to-neuron connectivity patterns, but with limited throughput. Barcode-based connectomics maps large numbers of single-neuron projections, but remains a challenge for jointly analyzing single-cell transcriptomics. Here, we established a rAAV2-retro barcode-based multiplexed tracing method that simultaneously characterizes the projectome and transcriptome at the single neuron level. We uncovered dedicated and collateral projection patterns of ventromedial prefrontal cortex (vmPFC) neurons to five downstream targets and found that projection-defined vmPFC neurons are molecularly heterogeneous. We identified transcriptional signatures of projection-specific vmPFC neurons, and verified Pou3f1 as a marker gene enriched in neurons projecting to the lateral hypothalamus, denoting a distinct subset with collateral projections to both dorsomedial striatum and lateral hypothalamus. In summary, we have developed a new multiplexed technique whose paired connectome and gene expression data can help reveal organizational principles that form neural circuits and process information.
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