F‐actin distribution at nodes of Ranvier and Schmidt‐Lanterman incisures in mammalian sciatic nerves

Abstract

Very little is known about the function of the F-actin cytoskeleton in the regeneration and pathology of peripheral nerve fibers. The actin cytoskeleton has been associated with maintenance of tissue structure, transmission of traction and contraction forces, and an involvement in cell motility. Therefore, the state of the actin cytoskeleton strongly influences the mechanical properties of cells and intracellular transport therein. In this work, we analyze the distribution of F-actin at Schmidt-Lanterman Incisures (SLI) and nodes of Ranvier (NR) domains in normal, regenerating and pathologic Trembler J (TrJ/+) sciatic nerve fibers, of rats and mice. F-actin was quantified and it was found increased in TrJ/+, both in SLI and NR. However, SLI and NR of regenerating rat sciatic nerve did not show significant differences in F-actin, as compared with normal nerves. Cytochalasin-D and Latrunculin-A were used to disrupt the F-actin network in normal and regenerating rat sciatic nerve fibers. Both drugs disrupt F-actin, but in different ways. Cytochalasin-D did not disrupt Schwann cell (SC) F-actin at the NR. Latrunculin-A did not disrupt F-actin at the boundary region between SC and axon at the NR domain. We surmise that the rearrangement of F-actin in neurological disorders, as presented here, is an important feature of TrJ/+ pathology as a Charcot-Marie-Tooth (CMT) model.

Poly(ADP-ribosylation) is present in murine sciatic nerve fibers and is altered in a Charcot-Marie-Tooth-1E neurodegenerative model

“…To sum up, the evidence proves the PAR identity of IHF PAR signals, validating the biological findings, namely: (1) PAR colocalizes with with F-actin at non-compact myelin regions; (2) in a Charcot-Marie-Tooth demyelinating disease model in which sciatic nerves present an excess of F-actin, accompanied by E-cadherin delocalization (Kun et al, 2012a; Devaux & Scherer, 2005), detected PAR was excessive and was delocalized too; (3) nuclear PAR was detectable in WT and moderately increased in Tr-J sciatic nerve Schwann cells.…”

“…2G). Subsequently, a previously established structural criteria (Kun et al, 2012a), stating that roughly the inner 40% of the fiber diameter corresponds to the axon whereas the remnant outer region corresponds to the Schwann cell, was adopted. This assumption allowed to affirm that both in Schwann cells (Fig.…”

“…Male 70 to 90 days old (P70-P90) wild-type (WT) and heterozygous mice carrying a mutation in pmp-22 (Tr-J) from Jackson Laboratory (strain B6.D2-Pmp22 Tr-J/J) were killed by cervical dislocation. Sciatic nerve dissection was promptly carried out in less than one minute and followed by fixation through immersion in cold freshly prepared 3% w/v paraformaldehyde (PFA) in PHEM buffer (60 mM Pipes, 25 mM Hepes, 10 mM EGTA, 2 mM magnesium chloride, pH 7.2-7.6) for 1 h. This procedure is known to grant a faster fixation than systemic descendent perfusion in the singular case of mice sciatic nerves (Kun et al, 2012a; Kun et al, 2012b). Then, nerves were cryoprotected in sucrose/PHEM at 4 °C (increasing concentrations along 24 h: 5% to 30% w/v) (Kun et al, 2012a; Kun et al, 2012b).…”

“…In each experiment, the average relative intensity of actin and PAR signals along the fiber diameter was calculated. Besides, assuming that the intervals from 0 to 30% and from 70 to 100% correspond to Schwann cell (SC) regions whereas the range of 30 to 70% corresponds to axon regions (Kun et al, 2012a), it was possible to calculate the average relative intensity of F-actin and PAR in the axon. Then the average relative intensities of PAR and actin in the axon were assigned the value 1 and used to normalize all the measurements.…”

“…Tr-J mice display an increased number of Schwann cells (Robertson et al, 1997) as well as structural alterations in Schmidt Lanterman incisures and paranodes of their sciatic nerve fibers involving changes in autotypic adherens junctions components such as E-cadherin (Devaux & Scherer, 2005). Moreover, actin microfilaments abundance and distribution in sciatic nerve fibers are distorted (Kun et al, 2012a). …”