Three‐dimensional reconstruction of the cranial and anterior spinal nerves in early tadpoles of Xenopus laevis (Pipidae, Anura)

Abstract

Xenopus laevis is one of the most widely used model organism in neurobiology. It is therefore surprising, that no detailed and complete description of the cranial nerves exists for this species. Using classical histological sectioning in combination with fluorescent whole mount antibody staining and micro-computed tomography we prepared a detailed innervation map and a freely-rotatable three-dimensional (3D) model of the cranial nerves and anterior-most spinal nerves of early X. laevis tadpoles. Our results confirm earlier descriptions of the pre-otic cranial nerves and present the first detailed description of the post-otic cranial nerves. Tracing the innervation, we found two previously undescribed head muscles (the processo-articularis and diaphragmatico-branchialis muscles) in X. laevis. Data on the cranial nerve morphology of tadpoles are scarce, and only one other species (Discoglossus pictus) has been described in great detail. A comparison of Xenopus and Discoglossus reveals a relatively conserved pattern of the post-otic and a more variable morphology of the pre-otic cranial nerves. Furthermore, the innervation map and the 3D models presented here can serve as an easily accessible basis to identify alterations of the innervation produced by experimental studies such as genetic gain- and loss of function experiments.

FoxN3 is necessary for the development of the interatrial septum, the ventricular trabeculae and the muscles at the head/trunk interface in the African clawed frog, Xenopus laevis (Lissamphibia: Anura: Pipidae)

“…Another muscle, the diaphragmatico‐branchialis, was described recently and has not been considered in previous studies on FoxN3 in X. laevis . This muscle develops as late as the cucullaris muscle (personal observations), is innervated by the vagal nerve, and, like the cucullaris muscle, spans the head/trunk interface . The diaphragmatico‐branchialis is malformed in FoxN3 morphants, indicating a head origin of this muscle.…”

“…All incubations were carried out at room temperature (21°C to 25°C). Antibody staining was conducted on whole mounts according to standard protocols . After bleaching, specimens were washed three times in Phosphate Buffered Saline (PBS) + 0.1% Triton X‐100 for 20 minutes each.…”

“…This muscle develops as late as the cucullaris muscle (personal observations), is innervated by the vagal nerve, and, like the cucullaris muscle, spans the head/trunk interface. 43 The diaphragmatico-branchialis is malformed in FoxN3 morphants, indicating a head origin of this muscle. The observation that the cucullaris and diaphragmaticobranchialis muscles of X. laevis are governed by the same developmental mechanisms as the head musculature implies a head origin of both muscles.…”

“…Antibody staining was conducted on whole mounts according to standard protocols. 43,64 After bleaching, specimens were washed three times in Phosphate Buffered Saline (PBS) + 0.1% Triton X-100 for 20 minutes each. They were then blocked for two hours in DAKO antibody diluent.…”

The morphology, development, and evolution of the head of fishes: Foundational studies for a renaissance of comparative morphology

“…(a) Siberian sturgeon Acipenser baerii at developmental stage 43 (17 mm TL), muscles shown in tones of red, cartilage shown in tones of blue, nerves shown in tones of yellow; from Warth, Hilton, Naumann, Olsson, and Konstantinidis ( Figure a). (b) Clawed frog Xenopus laevis at developmental stage 47/48, nerves shown in various colours, brain shown in yellow, eye shown in white, cartilage shown in transparent blue; from Naumann and Olsson (: Figure a)…”