Ivo Vanický

A Simple and Reproducible Model of Spinal Cord Injury Induced by Epidural Balloon Inflation in the Rat

This paper describes a modification of a balloon-compression technique to produce spinal cord injury in adult rats. A 2-French Fogarty catheter is inserted into the dorsal epidural space through a small hole made in T10 vertebral arch, advanced cranially to T8-9 spinal level, and inflated for 5 min. Spinal cord damage is graded by increasing the volume of saline used to inflate the balloon. Quantitative neurological and histopathological outcomes are presented with three different volumes (10, 15, and 20 microl of saline) to characterize the gradation of injury. Volume of 15 microl produced complete paraplegia followed by gradual recovery, finally reaching approximately the middle of the scale used to quantitate the locomotor performance. With these animals, after 4 weeks, the center of the lesion shows complete loss of grey matter and partial sparing of the white matter. We conclude that 15 microl volume produced submaximal injury that will be useful for studying the pathophysiology and effects of protective therapies with this compression-injury model.

Localization of NADPHd-exhibiting neurons in the spinal cord of the rabbit

Segmental and laminar distributions of nicotinamide adenine dinucleotide phosphate diaphorase (NADPHd)‐exhibiting neurons were examined in the rabbit spinal cord by using horizontal, sagittal, and transverse sections. A large number of NADPHd‐positive neurons in the spinal cord of rabbit appeared to fall into six categories (N1–N6), but others could not be classified. Major cell groups of NADPHd‐exhibiting neurons were identified in the superficial dorsal horn and around the central canal at all spinal levels and in the intermediolateral cell column at thoracic and upper lumbar levels. NADPHd‐exhibiting neurons of the pericentral region were divided into a thin subependymal cell column containing longitudinally arranged, small bipolar neurons with processes penetrating deeply into the intermediolateral cell column and/or running rostrocaudally in the subependymal layer. The second pericentral cell column located more laterally in lamina X contains large, intensely stained NADPHd‐exhibiting neurons with long dendrites radiating in the transverse plane. In the pericentral region (lamina X), close association of NADPHd‐exhibiting somata and fibers and mostly longitudinally oriented blood vessels were detected. Neurons of the sacral parasympathetic nucleus, seen in segments S1–S3, exhibited prominent NADPHd cellular staining accompanied by heavily stained fibers extending from Lissauers tract through lamina I along the lateral edge of the dorsal horn to lamina V. A massive dorsal gray commissure, highly positive in NADPHd staining, was found in segments S1–S3. Scattered positive cells were also found in the deeper dorsal horn, ventral horn, and white matter. Fiberlike NADPHd staining was found in the superficial dorsal horn and pericentral region in all the segments studied. Dense, punctate, nonsomatic NADPHd staining was detected in the superficial dorsal horn, in the pericentral region all along the rostrocaudal axis, and in the nucleus phrenicus (segments C4–C5), nucleus dorsalis (segments Th2–L2), Onufs nucleus (segments S1–S3), and the dorsal part of the dorsal gray commissure (S1–S3). J. Comp. Neurol. 406:263–284, 1999.

Reduced nicotinamide adenine dinucleotide phosphate diaphorase in the spinal cord of dogs

The distribution of somatic, fibre-like and punctate, non-somatic reduced nicotinamide adenine dinucleotide phosphate (NADPH) diaphorase activity was examined in dog spinal cord using horizontal, sagittal and transverse sections. The morphological features of NADPH diaphorase exhibiting neurons divided into six different neuronal types (N1-N6) were described and their laminar distribution specified. Major cell groups were identified in the superficial dorsal horn and around the central canal at all spinal levels, and in the intermediolateral cell column at thoracic level. NADPH diaphorase exhibiting neurons of the pericentral region were distributed in a thin subependymal cell column containing longitudinally-arranged small bipolar neurons with processes penetrating deeply into the intermediolateral cell column and/or running rostrocaudally in the subependymal layer. The second pericentral cell column located more laterally in lamina X contains large, intensely-stained NADPH diaphorase exhibiting neurons with long dendrites radiating in the transverse plane. Neurons of the sacral parasympathetic nucleus seen in segments S1-S3 exhibited prominent NADPH diaphorase activity accompanied by heavily-stained fibres extending from Lissauers tract through lamina I along the lateral edge of the dorsal horn to lamina V. A massive dorsal gray commissure, with high NADPH diaphorase activity, was found in segments S1-S3. At the same segmental level a prominent group of moderately-stained motoneurons was detected in the dorsolateral portion of the anterior horn. Fibre-like NADPH diaphorase activity was found in the superficial dorsal horn and pericentral region in all segments studied. Punctate, non-somatic NADPH diaphorase activity was detected in the superficial dorsal horn, in the pericentral region all along the rostrocaudal axis and in the nucleus phrenicus (segments C4-C5), nucleus dorsalis (segments Th2-L2), nucleus Y (segments S1-S3), and the dorsal part of the dorsal gray commissure (S1-S3). A schematic diagram documenting the segmental and laminar distribution of NADPH diaphorase activity is given.

Limited minocycline neuroprotection after balloon-compression spinal cord injury in the rat

Minocycline (MC), a second-generation tetracycline and anti-inflammatory agent reportedly provides neuroprotection following CNS injury. The objective of this study was to examine the neuroprotective effects of short and long-term MC treatment using balloon-compression spinal cord injury (SCI) in the rat. Rats subjected to SCI were treated with MC for 1 day (1DMC group; total dose 180 mg/kg) or 5 days (5DMC group; total dose 450 mg/kg) or placebo. The effects of MC treatment on locomotor recovery (BBB scale) and spinal cord white and gray matter sparing were evaluated for up to 28 days. Morphometric analysis showed that while MC treatment spared spinal cord white and gray matter rostral to the lesion epicenter in both, 1DMC and 5DMC groups, sparing of white and gray matter areas was not observed caudal to the traumatic lesion. In addition, MC treatment had no effect on final locomotor recovery. Limited improvement of spinal cord post-compression consequences raises questions about the neuroprotection efficiency of MC treatment following compression SCI in the rat.

Immunosuppressant FK506: focusing on neuroprotective effects following brain and spinal cord injury.

The secondary damage that follows central nervous system (CNS) injury is a target for neuroprotective agents aimed at tissue and function sparing. FK506, a clinically used immunosuppressant, acts neuroprotectively in rat models of brain and spinal cord injury and ischemia. Evidence of in vivo experimental studies highlights the neuroprotective role of FK506 by its direct impact on various cell populations within the CNS. The participation of FK506 in modulation of post-traumatic inflammatory processes is a further potential aspect involved in CNS neuroprotection. In this review we provide an overview of the current laboratory research focusing on the multiple effects of FK506 on neuroprotection following CNS injury.

Silver impregnability of ischemia-sensitive neocortical neurons after 15 minutes of cardiac arrest in the dog

SummaryThe development of postischemic neuronal argyrophilia and the subsequent fate of argyrophilic neurons were studied in dogs after 15 min of complete cerebral ischemia and survival varying from l h to 7 days. Histopathological examination of the vulnerable neocortical region was performed using the Nauta degeneration method, and the time course of cellular changes was described. Clear-cut neuronal argyrophilia was found to precede cell body shrinkage and gradual disintegration corresponding to selective neuronal death. To clarify this initial stage of neuronal impregnability, the samples from the animals surviving 8 h postarrest were stained with toluidine blue or processed for electron microscopy, and the distribution of argyrophilic cells was confirmed to be identical with that of hyperchromatic or electron-dense cells. On the other hand, infrequently observed “tissue infarctions” exhibited no silver affinity in spite of apparent cellular damage. These findings indicate that enhanced impregnability is related to cyto-chemical processes incidental to the phenomenon of “selective neuronal death”, which can be readily detected by the Nauta method.

Local transcutaneous cooling of the spinal cord in the rat: effects on long-term outcomes after compression spinal cord injury.

This study tested efficiency of a novel thermoelectric cooler for local transcutaneous spinal cord cooling. Spinal cord compression was made by epidural balloon inflation at T8-T9 level of the spinal cord. Experimental animals (n = 20) were divided into two groups. In the hypothermic group, local cooling started 25 min after spinal cord injury and lasted for 1 h with paravertebral temperature maintained at 28.5°C (±0.3). Normothermic group underwent identical procedures, but their temperature was maintained normothermic. The assessment of neurologic recovery was performed once a week during a 4 weeks survival period. After 4 weeks animals were sacrificed and the extent of the spinal cord lesion morphometrically evaluated. There were no statistically significant intergroup differences in BBB scores and preserved volumes of the spinal cord tissue. In consecutive cross-sectional areas, hypothermic animals had significantly more preserved white matter at the cranial periphery of the lesion. It was concluded that mild posttraumatic hypothermia (31.8°C) had some protective effect on tissue loss after spinal cord injury but this effect was not associated with functional improvement.

Neurodegeneration induced by reversed microdialysis of NMDA; a quantitative model for excitotoxicity in vivo.

This study characterizes a quantifiable in vivo model of excitotoxicity. In halothane anesthetized rats, microdialysis probe was implanted into somatosensory cortex/striatum and perfused by various concentrations (1, 10, 50 and 100 mmol/l) of N-methyl-d-aspartate (NMDA) for 20 min. After 24 h, histological quantification confirmed that NMDA produced a concentration-dependent excitotoxic lesion. With 10 mmol/l NMDA, coadministration of magnesium reduced significantly, and 2-amino-5-phosphonovalerate blocked completely the development of excitotoxic injury.

Audiogenic seizures after neck tourniquet-induced cerebral ischemia in the rat

Development of audiogenic seizures (AGS) and their correlation with neurodegeneration were studied after 7.5 min of whole-brain ischemia. One day post-ischemia, all animals became hyperreactive and responded to auditory stimulation by generalized seizures. Neuronal necrosis developed already 6 h post-ischemia in inferior colliculi, reticular thalamic nucleus and hippocampal hilar region. Repeated ischemia did not induce any neurological changes, suggesting that the neurological effects are consequences of selective neuronal injury.

Fos Protein Expression in Sacral Spinal Cord in Relation to Early Phase of Cauda Equina Syndrome in Dogs

Abstract1. The aim of the present study is to map the incipient phase of Fos expression in the sacral spinal cord neuronal pools of multiple cauda equina constrictions canine model.2. Fos-positive neurons were found bilaterally in the lateral portion of superficial dorsal horn layers (Laminae I–III) and along the lateral edge of the dorsal horn accompanied by the lateral collateral pathway, fibers of Lissauers tract, terminating at the sacral parasympathetic nucleus. Similarly, high Fos expression was detected in the ventral portion of the dorsal sacral commissure and in the dorsomedial portion of the anterior horns at S1–S3 segment level. Finally, a clearly expressed Fos-positivity was disclosed bilaterally in the neuropil of the nucleus Y in the anterior horn.3. Data from the present study show that continuous stimulation of the central fibers of sacral dorsal root ganglia neurons, i.e., fibers of sacral primary afferents, unlike those using various stimulations of the peripheral fibres offers an unusual pattern of Fos-like immunoreactivity.