Mitochondrial Dynamics in Visual Cortex Are Limited In Vivo and Not Affected by Axonal Structural Plasticity

Abstract

[5] are often localized far away from the soma, mitochondria are actively transported to these sites [6][7][8][9][10][11]. Also, the removal and degradation of mitochondria are tightly regulated [9,12,13], because dysfunctional mitochondria are a source of reactive oxygen species, which can damage the cell [14]. Deficits in mitochondrial trafficking have been proposed to contribute to the pathogenesis of Parkinson's disease, schizophrenia, amyotrophic lateral sclerosis, optic atrophy, and Alzheimer's disease [13,[15][16][17][18][19]. In neuronal cultures, about a third of mitochondria are motile, whereas the majority remains stationary for several days [8,20]. Activity-dependent mechanisms cause mitochondria to stop at synaptic sites [7,8,20,21], which affects synapse function and maintenance. Reducing mitochondrial content in dendrites decreases spine density [22,23], whereas increasing mitochondrial content or activity increases it [7]. These bidirectional interactions between synaptic activity and mitochondrial trafficking suggest that mitochondria may regulate synaptic plasticity. Here we investigated the dynamics of mitochondria in relation to axonal boutons of neocortical pyramidal neurons for the first time in vivo. We find that under these circumstances practically all mitochondria are stationary, both during development and in adulthood. In adult visual cortex, mitochondria are preferentially localized at putative boutons, where they remain for several days. Retinal-lesion-induced cortical plasticity increases turnover of putative boutons but leaves mitochondrial turnover unaffected. We conclude that in visual cortex in vivo, mitochondria are less dynamic than in vitro, and that structural plasticity does not affect mitochondrial dynamics. RESULTS AND DISCUSSION Few Motile Mitochondria in Axons of Pyramidal Neurons in Visual Cortex In VivoTo fluorescently label mitochondria and the neuronal structures in which they are localized, we performed in utero electroporation with two DNA constructs driving expression of mitomTurquoise2 and membrane-associated YFP. A cranial window was implanted once mice had reached the age of 8-10 weeks. This allowed us to visualize mitochondria in axonal arbors of layer 2/3 pyramidal neurons in V1 in vivo using two-photon microscopy (Figures 1A-1C; Movie S1). Two to three weeks after cranial window implantation, optical imaging of intrinsic imaging was performed to localize monocular V1. One week later, axonal branches in layer 1 and upper layer 2 were imaged using in vivo two-photon microscopy. We found that the density of mitochondria was 0.09 per mm ( Figure 1D). Surprisingly, the fraction of axonal mitochondria that were motile was only 0.83% ( Figure 1E), much lower than the 10%-30% previously observed in neuronal cultures [6][7][8][9][10]. We therefore asked whether the difference in mitochondrial motility was caused by the different experimental condition (in vivo versus in vitro) or by the difference in the age of the imaged neurons. We assessed mitochondrial motilit...

Local translation in neuronal compartments: how local is local?

“…If mitochondria power local translation, it is not clear if they function as compartmentalized entities serving specific translation compartments. The stabilization of mitochondrial localization at presynaptic sites and dendritic branch points during development and plasticity supports this view. Careful examination of such stable mitochondrial compartments that might power local translation is warranted in the future.…”

Mitostasis in Neurons: Maintaining Mitochondria in an Extended Cellular Architecture

“…On the one hand, a substantial body of studies has shown that about 10–20% of the mitochondria in axons near the neuronal soma are moving at any given time, even under conditions in which neuronal activity is either intrinsically absent (due to surgical isolation) or suppressed by anesthesia (Bilsland et al, 2010; Gilley et al, 2012; Misgeld et al, 2007; Sorbara et al, 2014; Takihara et al, 2015). However, some other studies have suggested that baseline mitochondrial flux in fully mature neurites would be low, with a steep developmental decline (Faits et al, 2016; Lewis et al, 2016; Smit-Rigter et al, 2016). However, a full appreciation of the effects of branched neuronal geometry on local flux rates reconciles these seemingly conflicting reports: In some systems measurements are possible in proximal axonal locations, where flux reflects the totality of moving mitochondria needed to supply the arbor, while in others, due to imaging constraints, only very distal parts of the arbor can be visualized.…”

“…Somewhat surprisingly, the majority of in vivo studies has failed to detect acute effects of activity (either spontaneous or imposed) and plasticity-inducing manipulations on bulk transport rates (Faits et al, 2016; Smit-Rigter et al, 2016). This questions the prevailing notion of neuronal activity as a major driver of baseline transport (but cf.…”

“…Thus, some of the motile mitochondria are designated to move anterograde, down the axon, while others prefer retrograde movement; what creates this distinction is unknown. Mitochondria in the stationary pool, in contrast, remain in place often for as long as the experimenter can image (Faits et al, 2016; Lewis et al, 2016; Smit-Rigter et al, 2016). The movement of mitochondria is mediated by a motor adaptor complex that attaches the anterograde kinesin-1 motor (Kif5B in mammals) and retrograde dynein motor to the mitochondrial outer membrane (Figure 3 A).…”

Regulation of mitochondrial dynamics in astrocytes: Mechanisms, consequences, and unknowns

“…Whether the low mobility of mitochondria reflects a difference in the preparation (organotypic cultures vs. in vivo) or a difference in the developmental age of the astrocytes (neonatal vs. adult) is an open question. However, others have observed that mitochondrial movement decreases in neurons in vivo (Lewis et al, 2016;Smit-Rigter et al, 2016). Using 2-photon in vivo imaging of mTurquise2-labeled mitochondria, Smit-Rigter and colleagues report that 1% of mitochondria are mobile in layer 2/3 pyramidal neuron axons (V1) in either adult FIG URE 1 Visualization of mitochondrial distribution in vivo.…”