FGF-2 is required to prevent astrogliosis in the facial nucleus after facial nerve injury and mechanical stimulation of denervated vibrissal muscles

Recently, we have shown that manual stimulation of paralyzed vibrissal muscles after facial-facial anastomosis reduced the poly-innervation of neuromuscular junctions and restored vibrissal whisking. Using gene knock outs, we found a differential dependence of manual stimulation effects on growth factors. Thus, insulin-like growth factor- 1 and brain-derived neurotrophic factor are required to underpin manual stimulation-mediated improvements, whereas FGF-2 is not. The lack of dependence on FGF-2 in mediating these peripheral effects prompted us to look centrally, i. e. within the facial nucleus where increased astrogliosis after facial-facial anastomosis follows synaptic stripping. We measured the intensity of Cy3-fluorescence after immunostaining for glial fibrillary acidic protein (GFAP) as an indirect indicator of synaptic coverage of axotomized neurons in the facial nucleus of mice lacking FGF-2 (FGF-2(-/-) mice). There was no difference in GFAP-Cy3-fluorescence (pixel number, gray value range 17-103) between intact wildtype mice (2.12 +/- 0.37x10(7)) and their intact FGF-2(-/-) counterparts (2.12 +/- 0.27x10(7)) nor after facial-facial anastomosis + handling (wildtype: 4.06 +/- 0.32x10(7); FGF-2(-/-) : 4.39 +/- 0.17x10(7)). However, after facial-facial anastomosis, GFAP-Cy3-fluorescence remained elevated in FGF-2(-/-) -animals (4.54 +/- 0.12x10(7)), whereas manual stimulation reduced the intensity of GFAP-immunofluorescence in wild type mice to values that were not significantly different from intact mice (2.63 +/- 0.39x10). We conclude that FGF-2 is not required to underpin the beneficial effects of manual stimulation at the neuro-muscular junction, but it is required to minimize astrogliosis in the brainstem and, by implication, restore synaptic coverage of recovering facial motoneurons

Manual stimulation of the whisker pad after hypoglossal–facial anastomosis (HFA) using a Y-tube conduit does not improve recovery of whisking function

Facial nerve injury is a common clinical trauma involving long-term functional deficits with facial asymmetry leading to associated psychological issues and social hardship. We have recently shown that repair by hypoglossal-facial or facial-facial nerve surgical end-to-end anastomosis and suture [hypoglossal-facial anastomosis (HFA) or facial-facial anastomosis (FFA)] results in collateral axonal branching, polyinnervation of neuromuscular junctions (NMJs) and poor function. We have also shown that another HFA repair procedure using an isogenic Y-tube (HFA + Y-tube) and involving a 10-mm gap reduces collateral axonal branching, but fails to reduce polyinnervation. Furthermore, we have previously demonstrated that manual stimulation (MS) of facial muscles after FFA or HFA reduces polyinnervation of NMJs and improves functional recovery. Here, we examined whether HFA + Y-tube and MS of the vibrissal muscles reduce polyinnervation and restore function. Isogenic Y-tubes were created using abdominal aortas. The proximal hypoglossal nerve was inserted into the long arm and sutured to its wall. The distal zygomatic and buccal facial nerve branches were inserted into the two short arms and likewise sutured to their walls. Manual stimulation involved gentle stroking of the vibrissal muscles by hand mimicking normal whisker movement. We evaluated vibrissal motor performance using video-based motion analysis, degree of collateral axonal branching using double retrograde labeling and the quality of NMJ reinnervation in target musculature using immunohistochemistry. MS after HFA + Y-tube reduced neither collateral branching, nor NMJ polyinnervation. Accordingly, it did not improve recovery of function. We conclude that application of MS after hypoglossal-facial nerve repair using an isogenic Y-tube is contraindicated: it does not lead to functional recovery but, rather, worsens it