Neuronal Death in the CNS Autonomic Control Center Comes Very Early after Cardiac Arrest and Is Not Significantly Attenuated by Prompt Hypothermic Treatment in Rats

Figures

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  Figure 1. Cresyl Violet (CV) staining in main regions of the rat brain of the sham operated (a,e,i,m,q,u), and CA operated (b–d, f–h, j–l, n–p, r–t, v–x) groups. CV stainability is gradually pale (damaged) with time in the CA groups. Note that damage is distinct at 2 days after CA. BMA, basomedial amygdaloid nucleus; CA1, cornu ammonis 1; Cpu, caudate putamen; PH, posterior hypothalamic area; Re, reuniens thalamic nucleus; S1, primary somatosensory cortex; SO, stratum oriens; SP, stratum pyramidale; SR, stratum radiatum. Scale bar: 100 µm.

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  Figure 2. CV staining in the myelencephalon (a–l) and intermediate zone of the spinal cord (a1–p1) of the sham operated (a,e,i,a1,e1,i1,m1), and CA operated (b–d, f–h, j–l, b1–d1, f1–h1, j1–l1, n1–p1) groups. In the CA operated groups, CV positive cells are severely damaged in the myelencephalon and intermediate zone of the spinal cord at 2 days post-CA. 10, dorsal motor nucleus of vagus; CC, central canal; CVL, caudoventrolateral reticular nucleus; Gi, gigantocellular reticular nucleus; Sol, nucleus of solitary tract; Scale bars: 100 µm.

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  Figure 3. NeuN immunohistochemical staining in the thalamus (a,a1) and myelencephalon (e,e1,i,i1,m,m1) in the sham group, and F-J B histofluorescence staining in the thalamus (b–d, b1–d1) and myelencephalon (f–h, f1–h1, j–l, j1–l1, n–p, n1–p1) of the CA operated (b–d, f–h, j–l, n–p) and CA/hypothermia operated (b1–d1, f1–h1, j1–l1, n1–p1) groups. In the thalamus, F-JB positive cells (arrows) are shown and in the reuniens thalamic nucleus (Re) from 1 day post-CA. In the myelencephalon, F-JB positive cells (arrows) are found in the gigantocellular reticular nucleus (Gi), caudoventrolateral reticular nucleus (CVL), solitary nucleus (Sol) and dorsal motor nucleus of vagus (10). From 12 h post-CA, hypothermia significantly reduces the number of F-JB positive cells only in the Gi at 12 h post-CA. Scale bar = 100 µm. (q) mean numbers of F-JB positive cells in the CA and CA/hypothermia groups (n = 9 per group, * p < 0.05, vs. each CA group). The bars indicate the means ± SEM.

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  Figure 4. NeuN immunohistochemical staining in the intermediate zone of the spinal cord of the sham operated (a,a1,e,e1,i,i1,m,m1) and F-J B histofluorescence staining in the intermediate zone of the spinal cord of the CA operated (b–d, f–h, j–l, n–p), and CA/hypothermia operated (b1–d1, f1–h1, j1–l1, n1–p1) groups. Detection of F-JB positive cells (arrows) starts from 12 h post-CA, showing that numbers of F-JB positive cells are different between spinal cord levels. Note that hypothermia significantly reduces numbers of F-JB positive cells in all the levels only at 12 h after CA. Scale bars = 100 µm. (q) mean numbers of F-JB positive cells in the CA and CA/hypothermia groups (n = 9 per group, * p < 0.05, vs. each CA group). The bars indicate the means ± SEM.

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  Figure 5. Iba-1 immunohistochemical staining in the thalamus (a–d) and myelencephalon (e–p) of the sham operated (a,a1,e,e1,i,i1,m,m1), CA operated (b–d, f–h, j–l, n–p), and CA/hypothermia operated (b1–d1, f1–h1, j1–l1, n1–p1) groups. Typical resting form of Iba-1 immunoreactive microglia are found in the sham operated group. After CA, Iba-1 immunoreactive microglia are morphologically activated, and Iba-1 immunoreactivity is increased with time after CA. Iba-1 immunoreactivity in the CA/hypothermia operated group is not significantly different comparted to that in the CA operated group. 10, dorsal motor nucleus of vagus; CVL, caudoventrolateral reticular nucleus; Gi, gigantocellular reticular nucleus; Re, reuniens thalamic nucleus; Sol, nucleus of solitary tract. Scale bars = 50 µm. (q) Quantitative analysis of Iba-1 immunoreactivity in the CA and CA/hypothermia groups (n = 9 per group). The bars indicate the means ± SEM.

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  Figure 6. Iba-1 immunohistochemical staining in the spinal cord of the sham operated (a,a1,e,e1,i,i1,m,m1), CA operated (b–d, f–h, j–l, n–p), and CA/hypothermia operated (b1–d1, f1–h1, j1–l1, n1–p1) groups. After CA, Iba-1 immunoreactivity is increased with time. Hypothermia significantly decreases microgliosis in all spinal levels only at 12 h after CA. Scale bars = 50 µm. (q) Quantitative analysis of Iba-1 immunoreactivity in the spinal cord in the CA and CA/hypothermia groups (n = 9 per group, * p < 0.05, vs. each CA group). The bars indicate the means ± SEM.

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  Figure 7. Schematic drawings of neuronal damage/death in brain and spinal cord regions related to autonomic control in the rat. Neuronal death at 12 h, 1 day and 2 days post-CA is marked by red, blue and green, respectively. Yellow indicates that neuronal death is not shown until 2 days post-CA, and white indicates that we did not investigate neuronal death after CA. Generally, the earliest neuronal death occurs in the myelencephalon and spinal cord.