Protein synthesis inhibitors delay transneuronal death in the piriform cortex of young adult rats

It has been demonstrated that apoptotic cell death is an active process that is dependent on RNA and protein synthesis. The question remains as to whether neuronal death in adult, mammalian brains can also be demonstrated in vivo to be dependent on protein synthesis. To address this question we have analysed transneuronal death in the piriform (olfactory) cortex. Following unilateral olfactory bulb ablation in young adult rats, layer IIa of the piriform cortex undergoes rapid degeneration, that commences 12 h after ablation and that is almost complete at 48 h. In order to block protein synthesis, three to six subcutaneous injections of the short acting protein synthesis inhibitor anisomycin, were given at 2 h intervals beginning just before the ablation of the olfactory bulb. In other cases a single injection of the long acting protein synthesis inhibitor emetine were made intracerebrally just before or after olfactory bulb ablation. The number of dying cells was then counted in sections through the rostrocaudal extent of the piriform cortex. Both anisomycin and emetine injections markedly reduced the number of pyknotic cells in layer IIa of the piriform cortex after olfactory bulb ablation. The effect of anisomycin was dose-dependent, near lethal doses leading to an almost complete absence of cell death (six injections of 100 mg/kg). As the doses of anisomycin were reduced, more dying cells were observed. Emetine was only effective at near lethal doses (10 mg/kg) and showed a greater capacity to reduce the levels of cell death when injected into structures near the piriform cortex (e.g., accumbens nucleus) than when injected into more distant structures. To further confirm that the cell death observed was due to apoptosis, we analysed sections by tunel staining to demonstrate DNA fragmentation. We found that tunel-positive cells were also always pyknotic, one of the landmarks of apoptosis. The appearance of pyknotic cells labelled by the tunel method demonstrated that the dying cells in the piriform cortex did indeed undergo apoptosis.     

Apoptosis in a neonatal rat model of cerebral hypoxia-ischemia

BACKGROUND and PURPOSE: The mechanisms of excitotoxic cell death in cerebral ischemia are poorly understood. In addition to necrosis, apoptotic cell death may occur. The purpose of this study was to determine whether an established model of cerebral hypoxia-ischemia in the neonatal rat demonstrates any features of apoptosis. METHODS: Seven-day-old neonatal rats underwent bilateral, permanent carotid ligation followed by 1 hour of hypoxia, and their brains were examined 1, 3, and 4 days after hypoxia-ischemia. The severity of ischemic damage was assessed in the dentate gyrus and frontotemporal cortex by light microscopy. Immunocytochemistry was performed to detect the cleavage of actin by caspases, a family of enzymes activated in apoptosis. Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) reactivity was examined in the cortical infarction bed and dentate gyrus. Neonatal rat brain DNA was run on agarose gel electrophoresis to detect DNA fragmentation. Ethidium bromide-staining and electron microscopy were used to determine whether apoptotic bodies, 1 of the hallmarks of apoptosis, were present. RESULTS: The frontotemporal cortex displayed evidence of infarction, and in most rats the dentate gyrus showed selective, delayed neuronal death. Immunocytochemistry demonstrated caspase-related cleavage of actin. TUNEL and DNA electrophoresis provided evidence of DNA fragmentation. Ethidium bromide-staining and electron microscopy confirmed the presence of chromatin condensation and apoptotic bodies. CONCLUSIONS: Features of apoptosis are present in the described model of cerebral hypoxia-ischemia. Apoptosis may represent a mode of ischemic cell death that could be the target of novel treatments that could potentially expand the therapeutic window for stroke.     

In-vitro Biological Study to Evaluate the Toxic Potentials of Fibrous Materials

Newborn Sprague-Dawley rats received a single dose of 2 Gy X-rays and were killed 6 hr later. Dying cells were characterized by extreme chromatin condensation and nuclear fragmentation. Dying cells were distributed in the primary and secondary germinal zones and in other brain regions. Among these latter, dying cells occurred in the cortical layers of the olfactory bulb, layers II-III and VIb of the neocortex, piriform and entorhinal cortex, stratum oriens and pyramidale of the hippocampus, striatum, thalamus, amygdala, brainstem, internal granular layer of the cerebellum, and cerebral and cerebellar white matter. Dying cells were immature cells, neurons and glial cells (including radial glia). In-situ labeling of nuclear DNA fragmentation identified individual cells bearing fragmented DNA. Since the number of cells stained with this method was larger than the number of dying cells, as revealed with current histological techniques, it is suggested that nuclear DNA fragmentation precedes chromatin condensation and nuclear fragmentation in X-ray-induced apoptosis. Furthermore, agarose gel electrophoresis of extracted DNA from irradiated brains showed a "ladder" pattern which is typical of internucleosomal DNA fragmentation and endonuclease activation.     

The effects of olfactory and somatosensory desensitization on Fos-like immunoreactivity in the brains of pup-exposed postpartum rats.

Fos-like immunoreactivity (fos-lir) was examined in sites within the "maternal circuit" in postpartum female rats that received various sensory desensitizations and were exposed to pups for 1 or 2 hr. Neither olfactory bulbectomy nor thelectomy (nipple removal) significantly reduced the fos-lir in the anterior medial preoptic area (MPOA), although reductions following bulbectomy in medial amygdala did occur. Peripherally induced hyposmia by ZnSo? reduced fos-lir in the olfactory structures (olfactory bulbs, piriform cortex, and olfactory tubercle), in medial and cortical nuclei of the amygdala, but not in anterior MPOA. Application of the topical anesthetic Emla to the ventrum only reduced fos-lir in the somatosensory cortex. Combined olfactory and ventral desensitizations produced marginal reductions in posterior MPOA. It is suggested that the MPOA is primarily involved as part of the effector system in the expression of the behavior. In contrast, the amygdala is involved in processing sensory cues received from pups during dam-litter interactions. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Cholinergic agonist carbachol enables associative long-term potentiation in piriform cortex slices

Pyramidal cells in piriform (olfactory) cortex receive afferent input from the olfactory bulb as well as intrinsic association input from piriform cortex and other cortical areas. These two functionally distinct inputs terminate on adjacent apical dendritic segments of the pyramidal cells located in layer Ia and layer Ib of piriform cortex. Studies with bath-applied cholinergic agonists have shown suppression of the fast component of the inhibitory postsynaptic potentials (IPSPs) evoked by stimulation of the association fibers. It was previously demonstrated that an associative form of LTP can be induced by coactivation of the two fiber systems after blockade of the fast, gamma-aminobutyric acid-A-mediated IPSP. In this report, we demonstrate that an associative form of long-term potentiation can be induced by coactivation of afferent and intrinsic fibers in the presence of the cholinergic agonist carbachol.     

Experimental brain injury induces regionally distinct apoptosis during the acute and delayed post-traumatic period

The temporal pattern of apoptosis in the adult rat brain after lateral fluid-percussion (FP) brain injury was characterized using terminal deoxynucleotidyl-transferase-mediated biotin-dUTP nick end labeling (TUNEL) histochemistry and agarose gel electrophoresis. Male Sprague Dawley rats were subjected to brain injury and killed for histological analysis at intervals from 12 hr to 2 months after injury (n = 3/time point). Sham (uninjured) controls were subjected to anesthesia with (n = 3) or without (n = 3) surgery. Apoptotic TUNEL-positive cells were defined using stringent morphological criteria including nuclear shrinkage and fragmentation and condensation of chromatin and cytoplasm. Double-labeled immunocytochemistry was performed to identify TUNEL-positive neurons (anti-neurofilament monoclonal antibody RM044), astrocytes (anti-glial fibrillary acidic protein polyclonal antibody), and oligodendrocytes (anti-cyclic nucleotide phosphohydrolase polyclonal antibody). Compared with that seen with sham controls, in the injured cortex, significant apoptosis occurred at 24 hr (65 +/- 19 cells; p < 0.05) with a second, more pronounced response at 1 week after injury (91 +/- 24 cells; p < 0.05). The number of apoptotic cells in the white matter was increased as early as 12 hr after injury and peaked by 1 week (33 +/- 6 cells; p < 0.05). An increase in apoptotic cells was observed in the hippocampus at 48 hr (13 +/- 8), whereas in the thalamus, the apoptotic response was delayed, peaking at 2 weeks after injury (151 +/- 71 cells; p < 0.05). By 2 months, the number of apoptotic cells in most regions had returned to uninjured levels. At 24 hr after injury, TUNEL-labeled neurons and oligodendrocytes were localized primarily to injured cortex. By 1 week after injury, populations of TUNEL-labeled astrocytes and oligodendrocytes were present in the injured cortex, while double-labeled neurons were present predominantly in injured cortex and thalamus, with a few scattered in the hippocampus. DNA agarose gels confirmed morphological identification of apoptosis. These data suggest that the apoptotic response to trauma is regionally distinct and may be involved in both acute and delayed patterns of cell death.     

Investigation of the tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone for in vivo and iIn vitro murine embryopathy and embryonic ras mutations

The evoked potential recorded in the rat piriform cortex in response to electrical stimulation of the olfactory bulb is composed of an early component occasionally followed by a late component (60-70 ms). We previously showed that the late component occurrence was enhanced following an olfactory learning. In the present study carried out in naive rats, we investigated the precise conditions of induction of this late component, and its spatiotemporal distribution along the olfactory pathways. In the anaesthetized rat, a stimulating electrode was implanted in the olfactory bulb. Four recording electrodes were positioned, respectively, in the olfactory bulb, the anterior and posterior parts of the piriform cortex, and the entorhinal cortex. Simultaneous recording of signals evoked in the four sampled structures in response to stimulation of the olfactory bulb revealed that the late component was detected in anterior and posterior piriform cortex as well as in entorhinal cortex, but not in the olfactory bulb. The late component occurred reliably for a narrow range of low intensities of stimulation delivered at frequencies not exceeding 1 Hz. Comparison of late component amplitude and latency across the different recorded sites showed that this component appeared first and with the greatest amplitude in the posterior piriform cortex. In addition to showing a functional dissociation between anterior and posterior parts of the piriform cortex, these data suggest that the posterior piriform cortex could be the locus of generation of this late high amplitude synchronized activity, which would then propagate to the neighbouring regions.     

Anatomical localization and time course of Fos expression following soman-induced seizures

Soman (pinacolymethylphosphonofluoridate), a highly potent, irreversible inhibitor of cholinesterase, causes intense convulsions, neuropathology and, ultimately, death. There is evidence that certain brain structures are selectively vulnerable to the pathological consequences of soman-induced seizures. A working hypothesis is that central nervous system (CNS) structures with the earliest and most severe signs of neuropathology may be key sites for the initiation of the seizures. Fos, the immediate-early gene product, increases rapidly in several animal seizure models. Thus, we reasoned that the earliest brain regions to express Fos might be involved in the initiation and maintenance of soman-induced convulsions. To assess this, rats were injected with a single, convulsive dose of soman (77.7 micrograms/kg, i.m.). The animals were euthanized and processed for immunocytochemical analysis at several time points. Robust Fos expression was seen in layer II of the piriform cortex and the noradrenergic nucleus locus coeruleus within 30-45 minutes. One hour following soman injection, staining was more intense in the piriform cortex layer II and in the locus coeruleus. In addition, Fos was evident in the piriform cortex layer III, the entorhinal cortex, the endopiriform nucleus, the olfactory tubercle, the anterior olfactory nucleus and the main olfactory bulb. By 2 hours, Fos staining was present throughout the cerebral cortex, thalamus, caudate-putamen and the hippocampus. At 8 hours and beyond, Fos expression returned to control levels throughout the CNS except for the piriform cortex and the locus coeruleus which still had robust labeling. By 24 hours, neuropathology was evident throughout the rostral-caudal extent of layer II of the piriform cortex. The rapid induction of Fos in the piriform cortex and the locus coeruleus, taken together with previous anatomical, eletrophysiological and neurochemical studies, suggests that prolonged, excessive exposure to synaptically released acetylcholine and norepinephrine triggers the production of soman-induced seizures initially in the piriform cortex and subsequently in other cortical and subcortical structures.     

Activation of CPP32-like caspases contributes to neuronal apoptosis and neurological dysfunction after traumatic brain injury

We examined the temporal profile of apoptosis after fluid percussion-induced traumatic brain injury (TBI) in rats and investigated the potential pathophysiological role of caspase-3-like proteases in this process. DNA fragmentation was observed in samples from injured cortex and hippocampus, but not from contralateral tissue, beginning 4 hr after TBI and continuing for at least 3 d. Double labeling of brain with terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) and an antibody directed to neuronal nuclear protein identified apoptotic neurons with high frequency in both traumatized rat cortex and hippocampus. Cytosolic extracts from injured cortex and hippocampus, but not from contralateral or control tissue, induced internucleosomal DNA fragmentation in isolated nuclei with temporal profiles consistent with those of DNA fragmentation observed in vivo. Caspase-3 mRNA levels, estimated by semiquantitative RT-PCR, were elevated fivefold in ipsilateral cortex and twofold in hippocampus by 24 hr after TBI. Caspase-1 mRNA content also was increased after trauma, but to a lesser extent in cortex. Increased caspase-3-like, but not caspase-1-like, enzymatic activity was found in cytosolic extracts from injured cortex. Intracerebroventricular administration of z-DEVD-fmk-a specific tetrapeptide inhibitor of caspase-3-before and after injury markedly reduced post-traumatic apoptosis, as demonstrated by DNA electrophoresis and TUNEL staining, and significantly improved neurological recovery. Together, these results implicate caspase-3-like proteases in neuronal apoptosis induced by TBI and suggest that the blockade of such caspases can reduce post-traumatic apoptosis and associated neurological dysfunction.     

Role of the net architecture in piriform cortex activity: analysis by a mathematical model

We present a mathematical analysis of the piriform cortex activity in rats. Experimental data were obtained by means of optical recording of fluorescent signals driven by neuronal activity. From these data, we determined the numerical value of the relaxation time for the pyramidal cell activity in layers II and III and the time latency map for bulb activation. Our model for the piriform cortex is based on pairs of excitatory and inhibitory neurons which correspond to pyramidal cells of layers II and III and to their inhibitory associated interneurons respectively; pyramidal cells are also interconnected through short and long range association fiber systems. Under such conditions, the model outputs resemble closely the experimental observations: (1) a double-bumped response to a strong and short stimulation; (2) oscillatory behavior under weak sustained stimulation conditions; (3) propagation of traveling activity waves; and (4) pacemaker activity when clusters of neurons are preferentially coupled.     

MR cholangiography in children after liver transplantation from living related donors

A highly specific anti-glutamate monoclonal antibody, mAb2D7, was used together with light and electron microscopy to elucidate the role played by the amino acid glutamate in the projection from the olfactory bulb to the piriform cortex in the rat. By light microscopy, glutamate-like immunoreactivity was observed in neuronal cell bodies and in the neuropil of the piriform cortex. Double labelling experiments which involved injections of wheat germ agglutinin-horse--radish peroxidase into the olfactory bulb and a post-embedding immunogold method for electron microscopy revealed anterogradely labelled terminals making asymmetric synaptic contacts on dendrites in the piriform cortex which contained high levels of glutamate as assessed by quantification. These results further support a role for glutamate as a neurotransmitter in the efferent pathway of the rat olfactory bulb.     

Beta-frequency (15-35 Hz) electroencephalogram activities elicited by toluene and electrical stimulation in the behaving rat

Bursts of beta-frequency (15-35 Hz) electroencephalogram activity occur in the olfactory system during odour sampling, but their mode of propagation within the olfactory system and potential contribution to the mechanisms of learning and memory are unclear. We have elicited large-amplitude beta activity in the rat olfactory system by applying noxious olfactory stimuli (toluene), and have monitored the bursts via chronically-implanted electrodes. Following exposure to toluene, coherent bursts with a peak frequency of 19.8 +/- 0.9 Hz were observed in the olfactory bulb, piriform cortex, entorhinal cortex and dentate gyrus. The timing of the bursts and the phases of electroencephalogram cross-spectra indicate that beta bursts propagate in a caudal direction from the olfactory bulb to the entorhinal cortex. The time delays between peaks of bursts in these structures were similar to latency differences for field potentials evoked by olfactory bulb or piriform cortex test-pulses. Peaks of burst cycles in the dentate region, however, were observed just prior to those in the entorhinal cortex. Surprisingly, power in toluene-induced beta-frequency oscillations was not increased following long-term potentiation induced by tetanic stimulation of the olfactory bulb, piriform cortex and entorhinal cortex. The activity of local inhibitory mechanisms may therefore counteract the effects of synaptic enhancements in afferent pathways during beta bursts. Low-frequency electrical stimulation of the piriform cortex was most effective in inducing coherent oscillatory responses in the entorhinal cortex and dentate gyrus at stimulation frequencies between 12 and 16 Hz. The results show that repetitive polysynaptic volleys at frequencies in the beta band induced by either toluene or electrical stimulation are transmitted readily within the olfactory system. The propagation of neural activity within this frequency range may therefore contribute to the transmission of olfactory signals to the hippocampal formation, particularly for those odours which induce high-amplitude bursts of beta activity.     

The HIV-1 gp120 causes ultrastructural changes typical of apoptosis in the rat cerebral cortex

We used transmission electron microscopy (TEM) and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick end-labelling (TUNEL) techniques to study the neuropathological effects of intracerebroventricular (i.c.v.) injection of recombinant HIV-1 gp 120 in rats. In brain cortical tissue sections from rats treated with a single daily dose of gp120 (100 ng day-1 for 7 or 14 consecutive days) TEM analysis showed chromatin compaction and marginalization along the inner surface of the nuclear envelope followed by masses of condensed chromatin, ultrastructural signs demonstrating the occurrence of apoptotic cell death. These effects were paralleled by in situ DNA fragmentation, as revealed by application of TUNEL technique to cryostat brain tissue sections from rats treated likewise with the viral coat protein. In no instance was apoptosis seen in the brain cortex of control rats. The present data demonstrate that gp120 given i.c.v. produces apoptosis in the neocortex of rats.     

ATP-sensitive voltage- and calcium-dependent chloride channels in sarcoplasmic reticulum vesicles from rabbit skeletal muscle

In addition to its role in olfaction and as a primary epileptogenic site, the anterior piriform cortex has been suggested to play a role in neuroperception of deficiencies or imbalances in physiologically essential amino acids. In recent studies, amino acid deficient diets were shown to induce expression of c-fos in the anterior piriform cortex within the rapid time frame associated with the normal anorectic response to such diets. It became important to examine the neurocytochemical architecture of this region for clues as to how and more precisely where dietary amino acid deficiency or imbalance might be monitored. The relationships of neuropeptide Y-, somatostatin-, and cholecystokinin-containing neurons were of particular interest because ongoing studies indicate that those peptides administered to the anterior piriform cortex alter intake of diets deficient in essential amino acids. The neuropeptides were endogenous to intrinsic neurons only; none resembled pyramidal projection neurons. Peptidergic neurons and fibers were concentrated most heavily in layer III of the paleocortex. The cytoarchitecture suggests that neuropeptide Y-, somatostatin-, and cholecystokin-containing neurons of the anterior piriform cortex may relate synaptically or multisynaptically to local circuit neurons during electrical activity, modulation of olfactory information, and neuroperception of essential amino acids.     

Olfactory-evoked regional cerebral blood flow in Alzheimer's disease.

Olfaction is impaired in Alzheimer's disease (AD). It was hypothesized that AD would reduce olfactory-evoked perfusion in mesial temporal olfactory (piriform) cortex, where neuropathology begins. Seven AD patients and 8 elderly controls (ECs) underwent olfactory threshold and identification tests and olfactory stimulation during positron emission tomography. Odor identification was impaired in AD, but threshold was not. Olfactory stimulation in ECs activated right and left piriform areas and right anterior ventral temporal cortex. AD patients had less activation in right piriform and anterior ventral temporal cortex but not in the left piriform area. Although orbital cortex did not activate in ECs, there was a significant between-groups difference in this area. Right piriform activation correlated with odor identification. Impaired odor identification likely reflects sensory cortex dysfunction rather than cognitive impairment. Given olfactory bulb projections to the mesial temporal lobe, olfactory stimulation during functional imaging might detect early dysfunction in this region. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Frequency-shaped amplification changes the neural representation of speech with noise-induced hearing loss

Apoptosis is a mode of active cell death. We have examined whether 2-chloroethylethyl sulfide (CEES), a sulfur vesicating agent, triggers apoptosis as a cytotoxic mechanism. Incubation of thymocytes with CEES, resulted in an induction of apoptotic features of cell death. Treatment of cells with 100 microM CEES for 5 h increased DNA fragmentation to approximately 40% of control. The fragmentation of DNA was visualized by agarose gel electrophoresis. It showed ladder pattern of DNA fragmentation, which indicates internucleosomal cleavage of DNA. Further evidence of apoptosis was observed in morphological changes of nuclei by using the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) method. The percentage of TUNEL positive cells was dependent upon CEES concentrations. CEES induced the classical morphological features of apoptosis in nucleus. These features were accompanied by condensation of chromatin, which arranged in sharply declined clumps and fragmentation of nucleus. To study requirement for synthesis of new protein in CEES-induced apoptosis, we studied the effect of cycloheximide for apoptotic activity. This protein synthesis inhibitor did not suppress the CEES-induced apoptotic activity. Taken together, these results suggest that CEES-induced apoptosis as a cytotoxicmechanism and this process occurs independent of synthesis of new protein.     

Noradrenergic suppression of synaptic transmission may influence cortical signal-to-noise ratio

Norepinephrine has been proposed to influence signal-to-noise ratio within cortical structures, but the exact cellular mechanisms underlying this influence have not been described in detail. Here we present data on a cellular effect of norepinephrine that could contribute to the influence on signal-to-noise ratio. In brain slice preparations of the rat piriform (olfactory) cortex, perfusion of norepinephrine causes a dose-dependent suppression of excitatory synaptic potentials in the layer containing synapses among pyramidal cells in the cortex (layer Ib), while having a weaker effect on synaptic potentials in the afferent fiber layer (layer Ia). Effects of norepinephrine were similar in dose-response characteristics and laminar selectivity to the effects of the cholinergic agonist carbachol, and combined perfusion of both agonists caused effects similar to an equivalent concentration of a single agonist. In a computational model of the piriform cortex, we have analyzed the effect of noradrenergic suppression of synaptic transmission on signal-to-noise ratio. The selective suppression of excitatory intrinsic connectivity decreases the background activity of modeled neurons relative to the activity of neurons receiving direct afferent input. This can be interpreted as an increase in signal-to-noise ratio, but the term noise does not accurately characterize activity dependent on the intrinsic spread of excitation, which would more accurately be described as interpretation or retrieval. Increases in levels of norepinephrine mediated by locus coeruleus activity appear to enhance the influence of extrinsic input on cortical representations, allowing a pulse of norepinephrine in an arousing context to mediate formation of memories with a strong influence of environmental variables.     

Evidence for a role of the chemorepellent semaphorin III and its receptor neuropilin-1 in the regeneration of primary olfactory axons

To explore a role for chemorepulsive axon guidance mechanisms in the regeneration of primary olfactory axons, we examined the expression of the chemorepellent semaphorin III (sema III), its receptor neuropilin-1, and collapsin response mediator protein-2 (CRMP-2) during regeneration of the olfactory system. In the intact olfactory system, neuropilin-1 and CRMP-2 mRNA expression define a distinct population of olfactory receptor neurons, corresponding to immature (B-50/GAP-43-positive) and a subset of mature (olfactory marker protein-positive) neurons located in the lower half of the olfactory epithelium. Sema III mRNA is expressed in pial sheet cells and in second-order olfactory neurons that are the target cells of neuropilin-1-positive primary olfactory axons. These data suggest that in the intact olfactory bulb sema III creates a molecular barrier, which helps restrict ingrowing olfactory axons to the nerve and glomerular layers of the bulb. Both axotomy of the primary olfactory nerve and bulbectomy induce the formation of new olfactory receptor neurons expressing neuropilin-1 and CRMP-2 mRNA. After axotomy, sema III mRNA is transiently induced in cells at the site of the lesion. These cells align regenerating bundles of olfactory axons. In contrast to the transient appearance of sema III-positive cells at the lesion site after axotomy, sema III-positive cells increase progressively after bulbectomy, apparently preventing regenerating neuropilin-1-positive nerve bundles from growing deeper into the lesion area. The presence of sema III in scar tissue and the concomitant expression of its receptor neuropilin-1 on regenerating olfactory axons suggests that semaphorin-mediated chemorepulsive signal transduction may contribute to the regenerative failure of these axons after bulbectomy.     

Neurotoxic effects of feline immunodeficiency virus, FIV-PPR

Apoptotic neuronal death is known to occur in the developing brain and in the mature brain of patients with ischemic and degenerative disorders. Although microglial cells are known to become activated in specific conditions, it has not been elucidated whether they enhance or prevent neuronal apoptosis. The present study was intended to observe how microglial cells are involved in neuronal death. When rat primary cortical neurons were incubated with a nitric oxide (NO) donor sodium nitroprusside (SNP; 300 microM) for 10 min, neuronal death occurred 12-16 hr later. The NO-induced neuronal death was inhibited by cycloheximide, and the SNP-treated neurons were characterized by nuclear fragmentation and intact cell membrane under electron microscopy. Agarose gel electrophoresis demonstrated DNA fragmentation of the SNP-treated neurons. Thus, the NO-induced neuronal death appeared to be apoptosis. When neurons were cocultured with rat primary microglial cells, the SNP treatment failed to induce the neuronal death. Because microglia-conditioned medium also prevented apoptotic neuronal death, microglial cells were considered to secrete antiapoptotic factors. The microglia-conditioned medium rescued neurons even when they were added to neuronal cultures after the SNP treatment, implying that the factors acted on neurons in a manner other than scavenging NO. Interleukin-3, interleukin-6, macrophage colony-stimulating factor, and basic fibroblast growth factor, which are known to be secreted by microglial cells, were not effective in preventing NO-induced neuronal death. Among microglia-derived substances, tumor necrosis factor alpha and plasminogen, which are heat-labile proteins, inhibited neuronal apoptosis. The neuroprotective action of the microglia-conditioned medium, however, still remained, even after it was heated. These findings suggest that microglial cells protect neurons against NO-induced lethal damage by secreting heat-labile and heat-stable neuroprotective factors in vitro.     

Spatiotemporal relationship of apoptotic cell death to lymphomonocytic infiltration in photochemically induced focal ischemia of the rat cerebral cortex

In this study we examined the time course of apoptotic cell death after photochemically induced focal ischemia of the rat cerebral cortex. For unequivocal differentiation between apoptosis and necrosis two criteria of programmed cell death were used: terminal deoxyribonucleotidyl transferase-mediated dUTP-digoxigenin nick end labeling (TUNEL) and morphological evidence of fragmentation and marginalization of nuclei. After photothrombosis, many TUNEL-positive cells were found within the infarct region from 12 h to 3 days. By day 6 they were preferentially located in the boundary zone of the infarct, and by day 14 they had disappeared. A high proportion of TUNEL-positive cells displayed fragmentation or marginalization of their nuclei, indicating apoptosis. Neurons, but not T cells and macrophages, were apoptotic. Inflammatory infiltrates were in close contact to apoptotic neurons throughout the infarct areas at day 1 and in the boundary zone between days 2 and 6 after photothrombosis. In summary, our study shows that neuronal apoptosis after cerebral ischemia is a prolonged process to which leukocyte-derived cytokines may contribute. In contrast to autoimmune diseases of the nervous system, termination of the local inflammatory response after cerebral ischemia does not involve apoptosis.