Abstract Spina bifida aperta (SBA), one of the most common congenital malformations, causes various neurological disorders. Pain is a common complaint of patients with SBA. However, little is known about the neuropathology of SBA-related pain. Because loss of γ-aminobutyric acid (GABA)ergic neurons in the spinal cord dorsal horn is associated with pain, we hypothesized the existence of cross-talk between SBA-related pain and alterations in GABAergic transmission in the spinal cord. Therefore, we investigated the kinetics of GABAergic transmission in the spinal cord dorsal horn in a chicken model of SBA. Neonatal chicks with SBA exhibited various pain-like behaviors, such as an increased number of vocalizations with elevated intensity (loudness) and frequency (pitch), reduced mobility, difficulty with locomotion, and escape reactions. Furthermore, the chicks with SBA did not respond to standard toe-pinching, indicating disruption of the spinal cord sensorimotor networks. These behavioral observations were concomitant with loss of GABAergic transmission in the spinal cord dorsal horn. We also found apoptosis of GABAergic neurons in the superficial dorsal horn in the early neonatal period, although cellular abnormalization and propagation of neurodegenerative signals were evident at middle to advanced gestational stages. In conclusion, ablation of GABAergic neurons induced alterations in spinal cord neuronal networks, providing novel insights into the pathophysiology of SBA-related pain-like complications.
Because excessive glutamate release is believed to play a pivotal role in numerous neuropathological disorders, such as ischemia or seizure, we aimed to investigate whether intrinsic prosaposin (PS), a neuroprotective factor when supplied exogenously in vivo or in vitro, is up-regulated after the excitotoxicity induced by kainic acid (KA), a glutamate analog. In the present study, PS immunoreactivity and its mRNA expression in the hippocampal and cortical neurons showed significant increases on day 3 after KA injection, and high PS levels were maintained even after 3 weeks. The increase in PS, but not saposins, detected by immunoblot analysis suggests that the increase in PS-like immunoreactivity after KA injection was not due to an increase in saposins as lysosomal enzymes after neuronal damage, but rather to an increase in PS as a neurotrophic factor to improve neuronal survival. Furthermore, several neurons with slender nuclei inside/outside of the pyramidal layer showed more intense PS mRNA expression than other pyramidal neurons. Based on the results from double immunostaining using anti-PS and anti-GABA antibodies, these neurons were shown to be GABAergic interneurons in the extra- and intra-pyramidal layers. In the cerebral cortex, several large neurons in the V layer showed very intense PS mRNA expression 3 days after KA injection. The choroid plexus showed intense PS mRNA expression even in the normal rat, and the intensity increased significantly after KA injection. The present study indicates that inhibitory interneurons as well as stimulated hippocampal pyramidal and cortical neurons synthesize PS for neuronal survival, and the choroid plexus is highly activated to synthesize PS, which may prevent neurons from excitotoxic neuronal damage. To the best of our knowledge, this is the first study that demonstrates axonal transport and increased production of neurotrophic factor PS after KA injection.
Prosaposin (PS) is a secretory neurotrophic factor, as well as a regulator of lysosomal enzymes. We previously reported the up-regulation of PS and the possibility of its axonal transport by GABAergic interneurons after exocitotoxicity induced by kainic acid (KA), a glutamate analog. In the present study, we performed double immunostaining with PS and three calcium binding protein markers: parvalbumin (PV), calbindin, and calretinin, for the subpopulation of GABAergic interneurons, and clarified that the increased PS around the hippocampal pyramidal neurons after KA injection existed mainly in the axons of PV positive interneurons. Electron microscopy revealed PS containing vesicles in the PV positive axon. Double immunostaining with PS and secretogranin or synapsin suggested that PS is secreted with secretogranin from synapses. Based on the results from in situ hybridization with two alternative splicing forms of PS mRNA, the increase of PS in the interneurons was due to the increase of PS + 0 (mRNA without 9-base insertion) as in the choroid plexus, but not PS + 9 (mRNA with 9-base insertion). These results were similar to those from the choroid plexus, which secretes an intact form PS + 0 to the cerebrospinal fluid. Neurons, especially PV positive GABAergic interneurons, produce and secrete the intact form of PS around hippocampal pyramidal neurons to protect them against KA neurotoxicity.
We examined the relationship between inherited motor-related conformation and orientation of facial hair whorls in Japanese Kiso horses. Eleven horses were divided into clockwise, counterclockwise, and radial groups according to facial hair whorls. We placed six markers on anatomical landmarks of each lateral side in a horse and measured the height of the landmarks, the distance between adjacent landmarks, and the angle of the adjacent landmarks. In the counterclockwise group, the horses tended to exhibit higher values on the left side than on the right side, and the comparison of the height of landmarks revealed a significant difference between both sides. Therefore, the orientation of facial hair whorls may suggest the tendency of motor-related conformation, at least in counterclockwise group.
Independent auditory end-organs appear first in amphibians in vertebrate phylogeny. In amphibians, sound detection is carried out by the amphibian papilla, basilar papilla and macula saccule. Amphibians inhabit distinct habitats and exhibit specific behaviours and sound frequency responses, so the amphibian vestibuloauditory system is an excellent model for considering the relationships between behaviour and physiological/anatomical vestibuloauditory properties. The African clawed frog, Xenopus laevis, lives in shallow water throughout its life and is thought to use sound in a higher frequency range compared with terrestrial anurans. In this study, the size of each vestibuloauditory end-organ and the distribution of ganglion cells in the vestibuloauditory ganglion were examined using haematoxylin and eosin staining and lectin histochemistry in Xenopus laevis. This study revealed that the size ratios among end-organs in Xenopus are similar to those in terrestrial anurans. Large and small cells were observed in the ganglion, but their distribution patterns are different from those in general terrestrial anurans. Lycopersicon esculentum lectin stained a large number of ganglion cells. Lectin-stained cells were found throughout the whole ganglion, but were especially abundant in the caudal part. These results suggested a unique distribution pattern of the vestibuloauditory ganglion cells in Xenopus.
