Evidence of the Primary Afferent Tracts Undergoing Neurodegeneration in Horses With Equine Degenerative Myeloencephalopathy Based on Calretinin Immunohistochemical Localization

Abstract

Equine degenerative myeloencephalopathy (EDM) is characterized by a symmetric general proprioceptive ataxia in young horses, and is likely underdiagnosed for 2 reasons: first, clinical signs overlap those of cervical vertebral compressive myelopathy; second, histologic lesions—including axonal spheroids in specific tracts of the somatosensory and motor systems—may be subtle. The purpose of this study was (1) to utilize immunohistochemical (IHC) markers to trace axons in the spinocuneocerebellar, dorsal column–medial lemniscal, and dorsospinocerebellar tracts in healthy horses and (2) to determine the IHC staining characteristics of the neurons and degenerated axons along the somatosensory tracts in EDM-affected horses. Examination of brain, spinal cord, and nerves was performed on 2 age-matched control horses, 3 EDM-affected horses, and 2 age-matched disease-control horses via IHC for calbindin, vesicular glutamate transporter 2, parvalbumin, calretinin, glutamic acid decarboxylase, and glial fibrillary acidic protein. Primary afferent axons of the spinocuneocerebellar, dorsal column–medial lemniscal, and dorsospinocerebellar tracts were successfully traced with calretinin. Calretinin-positive cell bodies were identified in a subset of neurons in the dorsal root ganglia, suggesting that calretinin IHC could be used to trace axonal projections from these cell bodies. Calretinin-immunoreactive spheroids were present in EDM-affected horses within the nuclei cuneatus medialis, cuneatus lateralis, and thoracicus. Neurons within those nuclei were calretinin negative. Cell bodies of degenerated axons in EDM-affected horses are likely located in the dorsal root ganglia. These findings support the role of sensory axonal degeneration in the pathogenesis of EDM and provide a method to highlight tracts with axonal spheroids to aid in the diagnosis of this neurodegenerative disease.

Transcriptome profiling of equine vitamin E deficient neuroaxonal dystrophy identifies upregulation of liver X receptor target genes

“…Even when frank cell death is evident at the histologic level, the site of selection for gene expression analyses in the CNS is often difficult to assess strictly based on anatomical data due to the diversity of cell types and their dependence on interconnectivity (59). Although lesions in eNAD occur at distal axons in the medulla oblongata (1), differential gene expression associated with the disease phenotype is most evident within the spinal cord, where the cell bodies for those particular axons reside (11). Gene expression studies in the Ttpa -null mouse have been previously conducted using microarray platforms (28, 29).…”

“…Based on our additional work determining that the nuclei for the affected neurons in eNAD originate in the dorsal root ganglia of the cervical spinal cord and terminate in the caudal medulla oblongata, (11) transcriptome sequencing was subsequently performed in the spinal cord, at the level of cervical vertebrae 1, in 5 eNAD-affected and 5-unaffected horses. For all samples, total RNA was extracted using TRIzol reagent (Thermofisher, Wilmington, DE, USA).…”

Concurrent Equine Degenerative Myeloencephalopathy and Equine Motor Neuron Disease in Three Young Horses

“…NAD/EDM‐affected horses have a general proprioceptive symmetric ataxia of all four limbs beginning at a few months of age whereas EMND‐affected horses have generalized lower motor neuron weakness, muscle atrophy, trembling, low head carriage, and tail head elevation, with a peak risk at 16 years of age . The histologic lesions associated with NAD/EDM are central axonal degeneration most pronounced in the somatosensory tracts (spinocuneocerebellar and dorsal spinocerebellar tract), with some involvement of the ventromedial motor tracts in more severely affected animals . Lesions associated with EMND include chromatolysis of neurons within the ventral columns as well as peripheral axonal degeneration and associated neurogenic atrophy of muscle fibers .…”

“…11 The histologic lesions associated with NAD/EDM are central axonal degeneration most pronounced in the somatosensory tracts (spinocuneocerebellar and dorsal spinocerebellar tract), with some involvement of the ventromedial motor tracts in more severely affected animals. 5,12 Lesions associated with EMND include chromatolysis of neurons within the ventral columns as well as peripheral axonal degeneration and associated neurogenic atrophy of muscle fibers. 6 Therefore, based on the age of onset, clinical signs and histologic lesions, NAD/EDM and EMND have always been categorized as distinct entities, despite the common association with an a-TP deficiency.…”

Equine degenerative myeloencephalopathy: prevalence, impact, and management

“… 2 Immunohistochemical calretinin staining has defined afferent proprioceptive tracts as the primary tracts damaged in eNAD/EDM horses, and isolated soma within the dorsal root ganglia as the source of dystrophic axons. 3 …”