DendritesIn VitroandIn VivoContain Microtubules of Opposite Polarity and Axon Formation Correlates with Uniform Plus-End-Out Microtubule Orientation

Abstract

In cultured vertebrate neurons, axons have a uniform arrangement of microtubules with plus-ends distal to the cell body (plus-end-out), whereas dendrites contain mixed polarity orientations with both plus-end-out and minus-end-out oriented microtubules. Rather than non-uniform microtubules, uniparallel minus-end-out microtubules are the signature of dendrites in Drosophila and Caenorhabditis elegans neurons. To determine whether mixed microtubule organization is a conserved feature of vertebrate dendrites, we used live-cell imaging to systematically analyze microtubule plus-end orientations in primary cultures of rat hippocampal and cortical neurons, dentate granule cells in mouse organotypic slices, and layer 2/3 pyramidal neurons in the somatosensory cortex of living mice. In vitro and in vivo, all microtubules had a plus-end-out orientation in axons, whereas microtubules in dendrites had mixed orientations. When dendritic microtubules were severed by laser-based microsurgery, we detected equal numbers of plus-and minus-end-out microtubule orientations throughout the dendritic processes. In dendrites, the minus-end-out microtubules were generally more stable and comparable with plus-end-out microtubules in axons. Interestingly, at early stages of neuronal development in nonpolarized cells, newly formed neurites already contained microtubules of opposite polarity, suggesting that the establishment of uniform plus-end-out microtubules occurs during axon formation. We propose a model in which the selective formation of uniform plus-end-out microtubules in the axon is a critical process underlying neuronal polarization.

Mechanisms and functions of lysosome positioning

“…In non-polarized cells, microtubule plus-ends are generally found in the peripheral cytoplasm and in neurons they point towards axon terminals, so, in these cells, kinesins mediate transport from the cell center towards the periphery (known as centrifugal transport). In some polarized cells, however, microtubules can point in other directions, as is the case for neuronal dendrites, which have microtubules with mixed orientations (Baas et al, 1988;Yau et al, 2016). In these cases, kinesins can potentially mediate both centrifugal and centripetal transport.…”

Microtubule nucleation and organization in dendrites

“…49 In the differentiating dendrite, microtubule nucleation and polymerization events occur away from the centrosome and throughout the growing arbor. 20,[27][28][29]35 Although microtubule nucleation events are spread throughout the arbor, specific foci repeatedly give rise to microtubules, suggesting that particular subcellular structures within the dendrite have microtubule nucleating activity. Other structures, in addition to the centrosome, can focus g-tubulin-mediated microtubule nucleation.…”

“…Immature differentiating dendrites have a larger balance of anterograde distal microtubules than mature dendrites and anterograde polymerization events are concentrated in the growing terminal regions. 17,[27][28][29]35 The polymerization of plus-end distal microtubules drives neurite outgrowth during neuron differentiation, 59 and live imaging of growing dendrites in vivo shows anterograde polymerizing microtubules extending the termini of emerging branches leading to outgrowth and stabilization 20,35 (Fig. 3A).…”

“…8,26 Both plus-or minus-end directed microtubule arrays are established in dendrites from the earliest stages of neurite outgrowth. 17,20,[27][28][29] One mechanism to achieve this organization is via the action of microtubule motor proteins. Motor proteins either anchored to the cell cortex or spanning two microtubules can facilitate the sliding of preassembled microtubules from the soma into the dendrite.…”

Microtubule Organization and Microtubule-Associated Proteins (MAPs)

“…This method also has limitations because it only detects dynamic MTs and not the stable MT population. Recently, in vivo imaging of growing MTs has confirmed the presence of a mixed MT organization in mature dendrites (Kleele et al 2014;Yau et al 2016). SHG microscopy allows label-free imaging without the addition of exogenous probes, but is hard to interpret quantitatively.…”