Biology subjects

Oertner, T.

Publications and source records attributed to Oertner, T..

3 recordsLinked to original sources

bioRxiv2019

Characterization of neuronal synaptopodin reveals a myosin V-dependent mechanism of synaptopodin clustering at the post-synaptic sites

The spine apparatus (SA) is an endoplasmic reticulum-related organelle which is present in a subset of dendritic spines in cortical and pyramidal neurons. The synaptopodin protein localizes between the stacks of the spine apparatus and is essential for the formation of this unique organelle. Although several studies have demonstrated the significance of the SA and synaptopodin in calcium homeostasis and plasticity of dendritic spines, it is still unclear what factors contribute to its stability at the synapse and whether the SA is locally formed or it is actively delivered to the spines. In this study we show that synaptopodin clusters are stable at their locations. We found no evidence of active microtubule-based transport for synaptopodin. Instead new clusters were emerging in the spines, which we interpret as the SA being assembled on-site. Furthermore, using super-resolution microscopy we show a tight association of synaptopodin with actin filaments. We identify the actin-based motor proteins myosin V and VI as novel interaction partners of synaptopodin and demonstrate that myosin V is important for the formation and/or maintenance of the SA.

neuroscience↗

bioRxiv2017

Ultrafast glutamate sensors resolve synaptic short-term plasticity

Glutamatergic synapses display a rich repertoire of plasticity mechanisms on many different time scales, involving dynamic changes in the efficacy of transmitter release as well as changes in the number and function of postsynaptic glutamate receptors. The genetically encoded glutamate sensor iGluSnFR enables visualization of glutamate release from presynaptic terminals at frequencies up to [~]10 Hz. However, to resolve glutamate dynamics during high frequency bursts, faster indicators are required. Here we report the development of fast (iGluf) and ultrafast (iGluu) variants with comparable brightness, but increased Kd for glutamate (137 M and 600 M, respectively). Compared to iGluSnFR, iGluu has a 6-fold faster dissociation rate in vitro and 5-fold faster kinetics in synapses. Fitting a three-state model to kinetic data, we identify the large conformational change after glutamate binding as the rate-limiting step. In rat hippocampal slice culture stimulated at 100 Hz, we find that iGluu is sufficiently fast to resolve individual glutamate release events, revealing that glutamate is rapidly cleared from the synaptic cleft. Depression of iGluu responses during 100 Hz trains correlates with depression of postsynaptic EPSPs, indicating that depression during high frequency stimulation is purely presynaptic in origin. At individual boutons, the recovery from depression could be predicted from the amount of glutamate released on the second pulse (paired pulse facilitation/depression), demonstrating differential frequency-dependent filtering of spike trains at Schaffer collateral boutons.\n\nSignificance StatementExcitatory synapses convert presynaptic action potentials into chemical signals that are sensed by postsynaptic glutamate receptors. To eavesdrop on synaptic transmission, genetically encoded fluorescent sensors for glutamate have been developed. However, even the best available sensors lag behind the very fast glutamate dynamics in the synaptic cleft. Here we report the development of an ultrafast genetically encoded glutamate sensor, iGluu, which allowed us to image glutamate clearance and synaptic depression during 100 Hz spike trains. We found that only boutons showing paired-pulse facilitation were able to rapidly recover from depression. Thus, presynaptic boutons act as frequency-specific filters to transmit select features of the spike train to specific postsynaptic cells.

neuroscience↗

bioRxiv2017

Artificial anion-conducting channelrhodopsins with tuned spectra, modified kinetics and enhanced light sensitivity

Genetic engineering of natural light-gated ion channels has proven a powerful way to generate optogenetic tools for a wide variety of applications. In recent years, blue light-activated engineered anion conducting channelrhodopsins (eACRs) have been developed, improved, and were successfully applied in vivo. We asked whether the approaches used to create eACRs can be transferred to other well-characterized cation-conducting channelrhodopsins (CCRs) to obtain eACRs with a broad spectrum of biophysical properties. We generated 22 variants using two conversion strategies applied to 11 CCRs and screened them for membrane expression, photocurrents and anion selectivity. We obtained two novel eACRs, Phobos and Aurora, with blue-and red-shifted action spectra and photocurrents similar to existing eACRs. Furthermore, step-function mutations greatly enhanced the cellular operational light sensitivity due to a slowed-down photocycle. These bistable eACRs can be reversibly toggled between open and closed states with brief light pulses of different wavelengths. All new eACRs reliably inhibited action potential firing in pyramidal CA1 neurons. In Drosophila larvae, eACRs conveyed robust and specific light-dependent inhibition of locomotion and nociception.

neuroscience↗