New study finds zebrafish have spinal enlargements linked to all fins, offering fresh insight into vertebrate evolution

Scientists from Nagoya University, Japan, have discovered that zebrafish possess enlarged areas in their spinal cords, previously believed to exist only in four-limbed vertebrates (tetrapods). The finding challenges long-standing assumptions about the evolution of spinal anatomy and its relationship to limb development.

Background: A Feature Once Thought Exclusive to Tetrapods

In tetrapods, the spinal cord has two enlarged regions corresponding to the forelimbs and hind limbs. These expansions are thought to have evolved to accommodate the complex muscle control and sensory input associated with limb movement.

Until now, scientists believed fish lacked such spinal enlargements, as they do not have true limbs.

Discovery: Subtle Enlargements in Zebrafish Spinal Cords

A research team led by Professor Naoyuki Yamamoto from the Graduate School of Bioagricultural Sciences at Nagoya University has now found that zebrafish do, in fact, possess spinal enlargements—although they are not visible to the naked eye.

“We thought that fish also have spinal enlargements because they have paired pectoral and pelvic fins, which correspond to forelimbs and hind limbs in tetrapods, respectively,” said Naoyuki Yamamoto, lead author of the study published in the journal Brain, Behavior and Evolution.

Methods: Mapping Fin-Related Spinal Regions

To test their hypothesis, the researchers examined which parts of the zebrafish spinal cord control the various paired and unpaired fins—the pectoral, pelvic, dorsal, caudal, and anal fins.

While previous studies had already identified the nerve connections for the pectoral, dorsal, and caudal fins, this research focused on the pelvic and anal fins.

The team used immunohistochemistry, a staining technique that highlights neurons, combined with a modified CUBIC (clear, unobstructed brain imaging cocktails) method. This approach allowed them to visualize deep spinal nerves that innervate the fins.

They then created serial tissue sections along the spinal cord to measure changes in the cross-sectional areas of both the spinal cord and the gray matter, correlating them with fin innervation levels.

Findings: Enlargements Detected Across All Fins

The analyses revealed that both the spinal cord and gray matter expanded in regions supplying nerves not only to paired fins (pectoral and pelvic) but also to unpaired fins (dorsal, anal, and caudal).

“We showed the presence of spinal enlargements in zebrafish, although they are modest and can only be detected through histological analysis,” Yamamoto stated. “Furthermore, we demonstrated that these enlargements are found in all fins—that is, both paired and unpaired fins.”

Evolutionary Implications: Rethinking the Origin of Limbs

The discovery suggests that spinal cord enlargements predate the evolution of limbs and were already present in the aquatic ancestors of tetrapods.

According to the researchers, when early fish evolved into land-dwelling tetrapods, only the paired fins—which were specialised for locomotion—transformed into limbs, while the unpaired fins disappeared.

This finding opens up new avenues for understanding how neural circuits adapted during the evolutionary transition from aquatic to terrestrial life.

Summary

• Zebrafish were found to have spinal enlargements, a feature once thought exclusive to four-limbed animals.
• Enlargements were detected in spinal regions connected to both paired and unpaired fins.
• The discovery was made using immunohistochemical staining and tissue section analysis.
• Findings suggest spinal enlargements evolved before limbs, during early vertebrate evolution.
• Research provides new insight into the neural and anatomical origins of limb control in vertebrates.
  

Original Publication
Authors: Ryo Takaoka, Hanako Hagio and Naoyuki Yamamoto.
Journal: Brain Behavior and Evolution
DOI: 10.1159/000548184
Method of Research: Experimental study
Subject of Research: Animals
Article Title: Identification of “spinal enlargements” correlating with paired and unpaired fins in zebrafish
Article Publication Date: 29-Aug-2025

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