Two specific tongue regions receive bilateral hypoglossal innervation: A study in neonatal rat pups
Introduction
We have previously reported axonal sprouting across the midline of the tongue from the intact side to the hypoglossal (XII) nerve-resected side in neonatal rats.1 Although the speed of axonal sprouting to the contralateral tongue remains uncertain, early onset of axonal sprouting within 24 h after a nerve insult still raises the possibility of residual nerve components of bilateral innervation of the XII nerve. Although the problem of postmortem fibre tracing comes from difficulties in constant visualization of finely arborized neural networks as judged from the relatively large number of unsuccessful cases, we re-examined XII innervation in neonatal rat tongues using a large number of normal newborn pups and many tongue preparations cut serially with shorter intervals. In addition, synaptophysin and vesicular acetylcholine transporter (VAChT), presynaptic terminal markers for the neuromuscular junction,2, 3, 4, 5 were immunohistochemically detected in the unilaterally XII nerve-resected tongues, because tongue movement is essential for suckling in developing rats.6, 7, 8 We performed quantitative analyses of the nerve terminals across the midline of the tongue in neonatal rats with suckling disturbance induced by unilateral resection of the XII nerve. In this study, we will discuss the functional role of bilateral XII innervation of the tongue.
Section snippets
Animals
Newborn Wistar rats (Japan SLC Inc., Shizuoka, Japan) of both sexes were used in this study. Postnatal day (P) 0 refers to the first 24 h after birth. All procedures were conducted in accordance with the National Institutes of Health Guide for the Care and Use of Laboratory Animals, and protocols were approved by our Institutional Animal Care and Use Committee. Every effort was made to minimize animal suffering and pain.
Postmortem neuronal tracing
Normal pups were euthanatized with sodium pentobarbital (100 mg/kg,
DiI-labelled XII nerve components across the midline of the tongue
The characteristics of the DiI-labelled XII nerve components in the neonatal rats’ tongues are summarized in Table 1. DiI application to the XII nerve stumps produced various stages of neural fibre labelling in neonatal rats, from unsuccessful cases without fine-labelled fibres to successful cases with finely arborized labelled fibres distributed throughout the left side of the tongue. In the unsuccessful cases, DiI labelling occurred only in thick fibre bundles in the posterior one-third of
Discussion
Postmortem unilateral DiI labelling of the XII nerve in the tongue of the normal neonatal rats and synaptophysin/VAChT immunohistochemistry of the tongue of the unilaterally XII nerve-resected neonatal rats demonstrated the presence of two specific tongue regions where nerve fibres extended to the contralateral side across the midline at the early neonatal stage just after birth. Furthermore, by postmortem bilateral labelling of the left (DiI) and right (4-Di-10-ASP) XII nerves, it became
Conclusions
We concluded that neonatal rats have two specific tongue regions receiving bilateral XII innervation, which allowed suckling in unilaterally XII nerve-resected neonatal rats.
Funding
None.
Conflict of interest
None declared.
Ethical approval
The protocol was approved by the Institutional Animal Care and Use Committee of Shinshu University (#190016).
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