TRPM7 is critical for short-term synaptic depression by regulating synaptic vesicle endocytosis
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Curated by eLife
Evaluation Summary:
Jiang et al. present a study on a novel role of the ion channel TRPM7 in the regulated release of neurotransmitters and hormones. Their analysis focused on TRPM7-deficient chromaffin cells and neurons, which they studied using electrophysiological and imaging techniques. The data indicate that TRPM7 is activated during the compensatory endocytotis process that follows secretory vesicle fusion. This TRPM7 activation increases Ca2+ influx into the cytosol during endocytosis, which boosts endocytosis and reduces the tendency of synapses to progressively fatigue in phases of prolonged stimulation. Multiple aspects of the present findings are novel, interesting, and important - they provide important new insights into the mechanisms by which transmitter and hormone release is fine-tuned. The mechanism of how secretory vesicles, and in particular synaptic vesicles, recycle is important to understand, as impairment in vesicle cycling in turn impairs synaptic transmission, and therefore brain function. However, there are several issues that remain to be addressed before the present data fully justify the conclusions put forward by the authors.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. The reviewers remained anonymous to the authors.)
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Abstract
Transient receptor potential melastatin 7 (TRPM7) contributes to a variety of physiological and pathological processes in many tissues and cells. With a widespread distribution in the nervous system, TRPM7 is involved in animal behaviors and neuronal death induced by ischemia. However, the physiological role of TRPM7 in central nervous system (CNS) neuron remains unclear. Here, we identify endocytic defects in neuroendocrine cells and neurons from TRPM7 knockout (KO) mice, indicating a role of TRPM7 in synaptic vesicle endocytosis. Our experiments further pinpoint the importance of TRPM7 as an ion channel in synaptic vesicle endocytosis. Ca 2+ imaging detects a defect in presynaptic Ca 2+ dynamics in TRPM7 KO neuron, suggesting an importance of Ca 2+ influx via TRPM7 in synaptic vesicle endocytosis. Moreover, the short-term depression is enhanced in both excitatory and inhibitory synaptic transmissions from TRPM7 KO mice. Taken together, our data suggests that Ca 2+ influx via TRPM7 may be critical for short-term plasticity of synaptic strength by regulating synaptic vesicle endocytosis in neurons.
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Author Response:
We appreciate reviewers’ favorable opinions regarding the significance and quality of our study, their excellent comments, and constructive criticisms. Following reviewers’ comments, we have performed additional experiments and re-analysis of our existing data, which confirms and strengthens conclusions of our study. We believe that we have been able to address all the reviewers’ concerns, and we thank the reviewers for the valuable comments, which have led to a substantial improvement of our manuscript.
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Evaluation Summary:
Jiang et al. present a study on a novel role of the ion channel TRPM7 in the regulated release of neurotransmitters and hormones. Their analysis focused on TRPM7-deficient chromaffin cells and neurons, which they studied using electrophysiological and imaging techniques. The data indicate that TRPM7 is activated during the compensatory endocytotis process that follows secretory vesicle fusion. This TRPM7 activation increases Ca2+ influx into the cytosol during endocytosis, which boosts endocytosis and reduces the tendency of synapses to progressively fatigue in phases of prolonged stimulation. Multiple aspects of the present findings are novel, interesting, and important - they provide important new insights into the mechanisms by which transmitter and hormone release is fine-tuned. The mechanism of how secretory …
Evaluation Summary:
Jiang et al. present a study on a novel role of the ion channel TRPM7 in the regulated release of neurotransmitters and hormones. Their analysis focused on TRPM7-deficient chromaffin cells and neurons, which they studied using electrophysiological and imaging techniques. The data indicate that TRPM7 is activated during the compensatory endocytotis process that follows secretory vesicle fusion. This TRPM7 activation increases Ca2+ influx into the cytosol during endocytosis, which boosts endocytosis and reduces the tendency of synapses to progressively fatigue in phases of prolonged stimulation. Multiple aspects of the present findings are novel, interesting, and important - they provide important new insights into the mechanisms by which transmitter and hormone release is fine-tuned. The mechanism of how secretory vesicles, and in particular synaptic vesicles, recycle is important to understand, as impairment in vesicle cycling in turn impairs synaptic transmission, and therefore brain function. However, there are several issues that remain to be addressed before the present data fully justify the conclusions put forward by the authors.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. The reviewers remained anonymous to the authors.)
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Joint Public Review:
The authors set out to assess the role of the ion channel TRPM7 in the regulated release of neurotransmitters and hormones. They used a TRPM7 KO mouse line and studied KO and WT control chromaffin cells and neurons using electrophysiological and imaging techniques. The basic finding is that TRPM7 seems to be activated during the endocytotic process that follows secretory vesicle fusion, and that this TRPM7 activation boosts Ca2+ influx into the cytosol upon endocytosis, resulting in augmentation of endocytosis. This is a quite unexpected discovery of a phenomenon that seems to have been missed in the many previous studies on chromaffin cell secretion - possibly due to the specific rectifying characteristics of TRPM7. The authors show further that TRPM7 KO synapses are characterized by less rapid synaptic depression, …
Joint Public Review:
The authors set out to assess the role of the ion channel TRPM7 in the regulated release of neurotransmitters and hormones. They used a TRPM7 KO mouse line and studied KO and WT control chromaffin cells and neurons using electrophysiological and imaging techniques. The basic finding is that TRPM7 seems to be activated during the endocytotic process that follows secretory vesicle fusion, and that this TRPM7 activation boosts Ca2+ influx into the cytosol upon endocytosis, resulting in augmentation of endocytosis. This is a quite unexpected discovery of a phenomenon that seems to have been missed in the many previous studies on chromaffin cell secretion - possibly due to the specific rectifying characteristics of TRPM7. The authors show further that TRPM7 KO synapses are characterized by less rapid synaptic depression, and link this phenomenon to altered endocytosis. Overall, key aspects of the data presented are convincing, novel, interesting, and important. On the other hand, there are multiple items that require clarification.
Strengths:
By examining the effects of TRPM7 loss in both chromaffin cells and dissociated hippocampal neurons, the authors are able to deploy a broad spectrum of techniques (each appropriate for the respective preparation) to dissect different aspects of the problem at hand.
Using cell attached membrane capacitance measurements, the authors show convincingly that when individual vesicles fuse, the time between exocytosis and endocytosis (i.e. membrane fission) is clearly increased upon TRPM7 deletion.
Amperometric analyses of secreted catecholamines show clearly that this delay in the fission time course does not impact the ability of vesicles to release cargo to the extracellular space.
The discovery that that the presence of TRPM7 allows for a divalent cation current to pass through the newly incorporated patch of membrane added to the plasma membrane during exocytosis is novel and interesting.
Deletion of TRPM7 slows down, by ~50%, synaptic vesicle endocytosis as assayed by either Synaptophysin-pHluorin or VGluT-pHluorin measurements in dissociated primary hippocampal neurons. This is a quite strong and hence striking effect.
The authors show convincingly that the impact of TRPM7 loss can be rescued by re-expressing a normal variant of the protein but not one that harbors a point-mutation that renders is non-conductive, indicating that ion flux through the channel may be required for the phenomenon analysed.
The authors show (using pHluorin-based measurements) that rapidly removing Ca2+ after a burst of exocytosis also slows down synaptic vesicle recycling kinetics - to the same extent as TRPM7 KO does. This indicates that the impact of removing external Ca is occluded in the KO background.
Weaknesses:
A basic analysis of the morphological properties and protein expression (e.g. of components of the endocytosis machinery) of TRPM7 KO chromaffin cells and neurons is missing.
The presentation of the results is often very brief, which hampers readability.
The authors did not thoroughly describe the rationale for several experimental strategy choices.
The experimental approach to test whether the slowed decay of the pHluorin signal is due to a slowing of the fission process or to a defect in vesicle reacidification was addressed, but the kinetics of the particular experiments are not very likely to resolve any differences even if they were there.
The authors report measurements of changes in nerve terminal intracellular Ca2+ based on readouts of Synptophysin-GCaMP6f in the KO versus the WT. These measurements reflect largely what is happening during a stimulus train, where there is minimal impact of the TRPM7 KO, and not what is happening during recovery (e.g. at the 60 s time point) - where according to the authors the presence of external Ca2+ and Ca2+ flux through TRPM7 is important.
There are some deficits with regard to linking the present experiments to what is already known about TRPM7.
Several experimental details need to be clarified and the discussion requires redaction, taking into account prior knowledge about TRPM7 and considering alternative explanations for some of the key findings.
The fact that TRPM7 loss slows down synaptic vesicle recycling and in turn alters synaptic depression would be of broad interest. However, there is no direct evidence that TRPM7 channels are present on synaptic vesicles, despite very extensive proteomics knowledge about the protein content of purified synaptic vesicles. Thus, it remains unclear whether and how the mechanism of a TRPM7-mediated Ca2+ influx is built into the secretory process, whether after exocytosis the channel appears on the plasma membrane, or if instead the channel is permamently present on the plasma membrane and is only activated - by as yet unknown mechanism - upon exocytosis.
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