Axons Of Peripheral Sensory Neurons Research Articles

Misregulation of mitochondria-lysosome contact dynamics in Charcot-Marie-Tooth Type 2B disease Rab7 mutant sensory peripheral neurons.

Inter-organelle contact sites between mitochondria and lysosomes mediate the crosstalk and bidirectional regulation of their dynamics in health and disease. However, mitochondria-lysosome contact sites and their misregulation have not been investigated in peripheral sensory neurons. Charcot-Marie-Tooth type 2B disease is an autosomal dominant axonal neuropathy affecting peripheral sensory neurons caused by mutations in the GTPase Rab7. Using live super-resolution and confocal time-lapse microscopy, we showed that mitochondria-lysosome contact sites dynamically form in the soma and axons of peripheral sensory neurons. Interestingly, Charcot-Marie-Tooth type 2B mutant Rab7 led to prolonged mitochondria-lysosome contact site tethering preferentially in the axons of peripheral sensory neurons, due to impaired Rab7 GTP hydrolysis-mediated contact site untethering. We further generated a Charcot-Marie-Tooth type 2B mutant Rab7 knock-in mouse model which exhibited prolonged axonal mitochondria-lysosome contact site tethering and defective downstream axonal mitochondrial dynamics due to impaired Rab7 GTP hydrolysis as well as fragmented mitochondria in the axon of the sciatic nerve. Importantly, mutant Rab7 mice further demonstrated preferential sensory behavioral abnormalities and neuropathy, highlighting an important role for mutant Rab7 in driving degeneration of peripheral sensory neurons. Together, this study identifies an important role for mitochondria-lysosome contact sites in the pathogenesis of peripheral neuropathy.

Target-Captured mRNA from Murine Olfactory Bulb.

The external world is perceived via sensory receptors arranged in highly organized systems according to functional strategies, which in turn reflect features of critical importance to both the sense and the animal. Thus describing the receptor organization and functional strategies of olfaction, the sense of smell, is crucial for a concrete understanding of the fundamental principles of the system's architecture. Sensory processing in olfactory systems is organized across olfactory bulb (OB) glomeruli, wherein axons of peripheral sensory neurons expressing the same olfactory receptor (OR) co-terminate to transmit receptor-specific activity to mitral/tufted cells, projection neurons in the olfactory bulb. Understanding how receptors map to glomeruli is therefore critical to understanding olfaction.Here we describe a method to enrich low-abundant OR mRNAs from the mouse OB for spatial transcriptomics to generate high-throughput mapping of receptors to glomeruli [2]. Our method combines sequential sectioning along the anteroposterior, dorsoventral, and mediolateral axes with target capture enrichment sequencing to overcome low-abundance target expression. This strategy spatially mapped 86% of olfactory receptors across the olfactory bulb and uncovered a relationship between olfactory receptor sequence and glomerular position.

Decoding the olfactory map through targeted transcriptomics links murine olfactory receptors to glomeruli

Sensory processing in olfactory systems is organized across olfactory bulb glomeruli, wherein axons of peripheral sensory neurons expressing the same olfactory receptor co-terminate to transmit receptor-specific activity to central neurons. Understanding how receptors map to glomeruli is therefore critical to understanding olfaction. High-throughput spatial transcriptomics is a rapidly advancing field, but low-abundance olfactory receptor expression within glomeruli has previously precluded high-throughput mapping of receptors to glomeruli in the mouse. Here we combined sequential sectioning along the anteroposterior, dorsoventral, and mediolateral axes with target capture enrichment sequencing to overcome low-abundance target expression. This strategy allowed us to spatially map 86% of olfactory receptors across the olfactory bulb and uncover a relationship between OR sequence and glomerular position.

Scales Radi(i)cally Remodel Sensory Axons and Vasculature

Peripheral axons of sensory neurons innervate skin cells to form a functional sensory organ. In this issue of Developmental Cell, Rasmussen etal. (2018) demonstrate that scale formation is essential for the development and regeneration of zebrafish sensory axons and vasculature.

Increased Expression of Transcription Factor SRY-box-Containing Gene 11 (Sox11) Enhances Neurite Growth by Regulating Neurotrophic Factor Responsiveness

The peripherally projecting axons of dorsal root ganglion (DRG) neurons readily regenerate after damage while their centrally projecting branches do not regenerate to the same degree after injury. One important reason for this inconsistency is the lack of pro-regeneration gene expression that occurs in DRG neurons after central injury relative to peripheral damage. The transcription factor SRY-box-containing gene 11 (Sox11) may be a crucial player in the regenerative capacity of axons as previous evidence has shown that it is highly upregulated after peripheral axon damage but not after central injury. Studies have also shown that overexpression or inhibition of Sox11 after peripheral nerve damage can promote or block axon regeneration, respectively. To further understand the mechanisms of how Sox11 regulates axon growth, we artificially overexpressed Sox11 in DRG neurons in vitro to determine if increased levels of this transcription factor could enhance neurite growth. We found that Sox11 overexpression significantly enhanced neurite branching in vitro, and specifically induced the expression of glial cell line-derived neurotrophic factor (GDNF) family receptors, GFRα1 and GFRα3. The upregulation of these receptors by Sox11 overproduction altered the neurite growth patterns of DRG neurons alone and in response to growth factors GDNF and artemin; ligands for GFRα1 and GFRα3, respectively. These data support the role of Sox11 to promote neurite growth by altering responsiveness of neurotrophic factors and may provide mechanistic insight as to why peripheral axons of sensory neurons readily regenerate after injury, but the central projections do not have an extensive regenerative capacity.

Intraneural Injection of ATP Stimulates Regeneration of Primary Sensory Axons in the Spinal Cord.

Injury to the peripheral axons of sensory neurons strongly enhances the regeneration of their central axons in the spinal cord. It remains unclear on what molecules that initiate such conditioning effect. Because ATP is released extracellularly by nerve and other tissue injury, we hypothesize that injection of ATP into a peripheral nerve might mimic the stimulatory effect of nerve injury on the regenerative state of the primary sensory neurons. We found that a single injection of 6 μl of 150 μm ATP into female rat sciatic nerve quadrupled the number of axons growing into a lesion epicenter in spinal cord after a concomitant dorsal column transection. A second boost ATP injection 1 week after the first one markedly reinforced the stimulatory effect of a single injection. Single ATP injection increased expression of phospho-STAT3 and GAP43, two markers of regenerative activity, in sensory neurons. Double ATP injections sustained the activation of phospho-STAT3 and GAP43, which may account for the marked axonal growth across the lesion epicenter. Similar studies performed on P2X7 or P2Y2 receptor knock-out mice indicate P2Y2 receptors are involved in the activation of STAT3 after ATP injection or conditioning lesion, whereas P2X7 receptors are not. Injection of ATP at 150 μm caused little Wallerian degeneration and behavioral tests showed no significant long-term adverse effects on sciatic nerve functions. The results in this study reveal possible mechanisms underlying the stimulation of regenerative programs and suggest a practical strategy for stimulating axonal regeneration following spinal cord injury.SIGNIFICANCE STATEMENT Injury of peripheral axons of sensory neurons has been known to strongly enhance the regeneration of their central axons in the spinal cord. In this study, we found that injection of ATP into a peripheral nerve can mimic the effect of peripheral nerve injury and significantly increase the number of sensory axons growing across lesion epicenter in the spinal cord. ATP injection increased expression of several markers for regenerative activity in sensory neurons, including phospho-STAT3 and GAP43. ATP injection did not cause significant long-term adverse effects on the functions of the injected nerve. These results may lead to clinically applicable strategies for enhancing neuronal responses that support regeneration of injured axons.

Charcot Marie Tooth 2B Peripheral Sensory Neuropathy: How Rab7 Mutations Impact NGF Signaling?

Charcot-Marie-Tooth 2B peripheral sensory neuropathy (CMT2B) is a debilitating autosomal dominant hereditary sensory neuropathy. Patients with this disease lose pain sensation and frequently need amputation. Axonal dysfunction and degeneration of peripheral sensory neurons is a major clinical manifestation of CMT2B. However, the cellular and molecular pathogenic mechanisms remain undefined. CMT2B is caused by missense point mutations (L129F, K157N, N161T/I, V162M) in Rab7 GTPase. Strong evidence suggests that the Rab7 mutation(s) enhances the cellular levels of activated Rab7 proteins, thus resulting in increased lysosomal activity and autophagy. As a consequence, trafficking and signaling of neurotrophic factors such as nerve growth factor (NGF) in the long axons of peripheral sensory neurons are particularly vulnerable to premature degradation. A “gain of toxicity” model has, thus, been proposed based on these observations. However, studies of fly photo-sensory neurons indicate that the Rab7 mutation(s) causes a “loss of function”, resulting in haploinsufficiency. In the review, we summarize experimental evidence for both hypotheses. We argue that better models (rodent animals and human neurons) of CMT2B are needed to precisely define the disease mechanisms.

Identification of possible downstream genes required for the extension of peripheral axons in primary sensory neurons

The LIM-homeodomain transcription factor Islet2a establishes neuronal identity in the developing nervous system. Our previous study showed that Islet2a function is crucial for extending peripheral axons of sensory neurons in zebrafish embryo. Overexpressing a dominant-negative form of Islet2a significantly reduced peripheral axon extension in zebrafish sensory neurons, implicating Islet2a in the gene regulation required for neurite formation or proper axon growth in developing sensory neurons. Based on this, we conducted systematic screening to isolate genes regulated by Islet2a and affecting the development of axon growth in embryonic zebrafish sensory neurons. The 26 genes selected included some encoding factors involved in neuronal differentiation, axon growth, cellular signaling, and structural integrity of neurons, as well as genes whose functions are not fully determined. We chose four representative candidates as possible Islet2a downstream functional targets (simplet, tppp, tusc5 and tmem59l) and analyzed their respective mRNA expressions in dominant-negative Islet2a-expressing embryos. They are not reported the involvement of axonal extension or their functions in neural cells. Finally, knockdown of these genes suggested their direct actual involvement in the extension of peripheral axons in sensory neurons.

Glial cell line-derived neurotrophic factor defines the path of developing and regenerating axons in the lateral line system of zebrafish

How the peripheral axons of sensory neurons are guided to distant target organs is not well understood. Here we examine this question in the case of the posterior lateral line (PLL) system of zebrafish, where sensory organs are deposited by a migrating primordium. Sensory neurites accompany this primordium during its migration and are thereby guided to their prospective target organs. We show that the inactivation of glial cell line-derived neurotrophic factor (GDNF) signaling leads to defects of innervation and that these defects are due to the inability of sensory axons to track the migrating primordium. GDNF signaling is also used as a guidance cue during axonal regeneration following nerve cut. We conclude that GDNF is a major determinant of directed neuritic growth and of target finding in this system, and we propose that GDNF acts by promoting local neurite outgrowth.

Developmentally Regulated Impediments to Skin Reinnervation by Injured Peripheral Sensory Axon Terminals

SummaryThe structural plasticity of neurites in the central nervous system (CNS) diminishes dramatically after initial development, but the peripheral nervous system (PNS) retains substantial plasticity into adulthood. Nevertheless, functional reinnervation by injured peripheral sensory neurons is often incomplete [1–6]. To investigate the developmental control of skin reinnervation we imaged the regeneration of trigeminal sensory axon terminals in live zebrafish larvae following laser axotomy. When axons were injured during early stages of outgrowth, regenerating and uninjured axons grew into denervated skin and competed with one another for territory. At later stages, after the establishment of peripheral arbor territories, the ability of uninjured neighbors to sprout diminished severely, and although injured axons reinitiated growth, they were repelled by denervated skin. Regenerating axons were repelled specifically by their former territories, suggesting that local inhibitory factors persist in these regions. Antagonizing the function of several members of the Nogo Receptor (NgR)/RhoA pathway improved the capacity of injured axons to grow into denervated skin. Thus, as in the CNS, impediments to reinnervation in the PNS arise after initial establishment of axon arbor structure.

Electrophysiological evidence for tetrodotoxin-resistant sodium channels in slowly conducting dural sensory fibers.

A tetrodotoxin (TTX)-resistant sodium channel was recently identified that is expressed only in small diameter neurons of peripheral sensory ganglia. The peripheral axons of sensory neurons appear to lack this channel, but its presence has not been investigated in peripheral nerve endings, the site of sensory transduction in vivo. We investigated the effect of TTX on mechanoresponsiveness in nerve endings of sensory neurons that innervate the intracranial dura. Because the degree of TTX resistance of axonal branches could potentially be affected by factors other than channel subtype, the neurons were also tested for sensitivity to lidocaine, which blocks both TTX-sensitive and TTX-resistant sodium channels. Single-unit activity was recorded from dural afferent neurons in the trigeminal ganglion of urethan-anesthetized rats. Response thresholds to mechanical stimulation of the dura were determined with von Frey monofilaments while exposing the dura to progressively increasing concentrations of TTX or lidocaine. Neurons with slowly conducting axons were relatively resistant to TTX. Application of 1 microM TTX produced complete suppression of mechanoresponsiveness in all (11/11) fast A-delta units [conduction velocity (c.v.) 5-18 m/s] but only 50% (5/10) of slow A-delta units (1.5 <c.v.<5 m/s) and 13% (2/15) of C units (c.v. </=1.5 m/s). The mean TTX concentration that produced complete suppression of mechanoresponsiveness was approximately 270-fold higher in C units than in fast A-delta units. In contrast, no significant difference was found between C and A-delta units in the concentration of lidocaine required for complete suppression of mechanoresponsiveness, indicating that the greater TTX resistance of mechanoresponsiveness in C units is not attributable to differences in safety factor unrelated to channel subtype. These data offer indirect evidence that a TTX-resistant channel subtype is expressed in the terminal axonal branches of many of the more slowly conducting (C and slow A-delta) dural afferents. The channel appears to be present in these fibers, but not in the faster A-delta fibers, in sufficient numbers to support the initiation and propagation of mechanically induced impulses. Comparison with previous data on the absence of TTX resistance in peripheral nerve fibers suggests that the TTX-resistant sodium channel may be a distinctive feature of the receptive rather than the conductive portion of the sensory neuron's axonal membrane.

Target-induced neurogenesis in the leech CNS involves efferent projections to the target

During a critical period in leech embryogenesis, the sex nerves that connect the 5th and 6th midbody ganglia (MG5 and MG6) to the primordium of the male sexual organ carry a spatially localized signal that induces the birth of several hundred neurons specific to these ganglia. We examined particular cellular elements (afferents, efferents, non-neuronal components) within these nerves as potential conveyors of the inductive signal. We show that axons of peripheral sensory neurons in the male genitalia travel along the sex nerves and into MG5 and MG6, but reach the CNS after the critical period has elapsed and cannot, therefore, be involved in the induction. Of the six sex nerves, four contain non-neuronal cells that span the entire distance between the male genitalia and the sex ganglia. However, when male genitalia were transplanted to ectopic locations close to MG6, induction occurred frequently but only in MG6, mediated by ectopic nerves that do not contain these cells. Thus, non-neuronal cells specific to the normal sex nerves are not necessary for induction. In addition, dye injections into the target during the critical period failed to reveal migrating cells in the sex nerves that could convey the inductive signal to the CNS. Finally, we show that 11 pairs of central neurons in each ganglion project to the male organ early during the critical period. In the adult, at least 3 additional pairs of neurons in MG6 also innervate this target. We conclude that the only components of the sex nerves that connect the sex ganglia to the target during the critical period that could be associated with induced central mitogenesis are the axons of central neurons that innervate the male genitalia.