Amanda McRae

Propentosylline depresses amyloid and Alzheimer's CSF microglial antigens after ischaemia.

In the gerbil hippocampus activated microglial antigens are intensely stained by cerebrospinal fluid from patients with Alzheimers disease (AD-CSF), OX18 and the amyloid precursor protein (beta-APP) up to 14 days after ischaemia. Propentosylline (PPF), which facilitates the adenosine A2 receptor action, has been shown to be neuroprotective, to depress O2- radical formation in macrophages and to interfere with the generation of phagocytotic macrophages from cultivated microglial cells. In this report we tested in ischaemic gerbils whether PPF treatment influences the potential neurotoxic properties of microglia. Daily post-treatment with PPF, started 24 h after ischaemia, depressed the immunostaining of activated microglia by AD-CSF, OX18 and APP in the hippocampus. Thus, PPF may protect against microglia-related brain damage.

Long-term potentiation protects rat hippocampal slices from the effects of acute hypoxia

We have previously shown that long-term potentiation (LTP) decreases the sensitivity of glutamate receptors in the rat hippocampal CA1 region to exogenously applied glutamate agonists. Since the pathophysiology of hypoxia/ischemia involves increased concentration of endogenous glutamate, we tested the hypothesis that LTP could reduce the effects of hypoxia in the hippocampal slice. The effects of LTP on hypoxia were measured by the changes in population spike potentials (PS) or field excitatory post-synaptic potentials (fepsps). Hypoxia was induced by perfusing the slice with (i) artificial CSF which had been pre-gassed with 95%N2/5% CO2; (ii) artificial CSF which had not been pre-gassed with 95% O2/5% CO2; or (iii) an oxygen-glucose deprived (OGD) medium which was similar to (ii) and in which the glucose had been replaced with sucrose. Exposure of a slice to a hypoxic medium for 1.5-3.0 min led to a decrease in the PS or fepsps; the potentials recovered to control levels within 3-5 min. Repeat exposure, 45 min later, of the same slice to the same hypoxic medium for the same duration as the first exposure caused a reduction in the potentials again; there were no significant differences between the degree of reduction caused by the first or second exposure for all three types of hypoxic media (P>0.05; paired t-test). In some of the slices, two episodes of LTP were induced 25 and 35 min after the first hypoxic exposure; this caused inhibition of reduction in potentials caused by the second hypoxic insult which was given at 45 min after the first; the differences in reduction in potentials were highly significant for all the hypoxic media used (P<0.01; paired t-test). The neuroprotective effects of LTP were not prevented by cyclothiazide or inhibitors of NO synthetase compounds that have been shown to be effective in blocking the effects of LTP on the actions of exogenously applied AMPA and NMDA, respectively. The neuroprotective effects of LTP were similar to those of propentofylline, a known neuroprotective compound. We conclude that LTP causes an appreciable protection of hippocampal slices to various models of acute hypoxia. This phenomenon does not appear to involve desensitisation of AMPA receptors or mediation by NO, but may account for the recognised inverse relationship between educational attainment and the development of dementia.

Propentofylline attenuates microglial reaction in the spinal cord induced by middle cerebral artery occlusion

This study examines the effect of Propentofylline (PPF) on reactive microglia in the lumbar spinal cord in rats following focal cerebral ischaemia produced by permanent occlusion of the middle cerebral artery (MCA). Our results showed that daily treatment of PPF beginning at 24 h after MCA occlusion for 2 or 4 consecutive days markedly suppressed the microglial response as detected immunohistochemically with OX-42. The most dramatic effect was the prevention of transformation of ramified microglia into amoeboidic form as well as formation of perineuronal microglia in close association with the soma of motoneurons. This has greatly amplified the potentiality of PPF used as a neuroprotective drug against microglia-related neuron damage induced by cerebral ischaemia.

Properties of activated microglia and pharmacologic interference by propentofylline.

Ameboid microglia are activated macrophages in the developing brain. With age, these cells undergo gradual transformation into the adult form, known as ramified or resting microglia. In response to neuronal insults, microglia change their morphology and immunophenotype and proliferate to become full-blown brain macrophages. Microglia release a battery of neurotoxic substances. Responses to neuronal damage occur at various intervals after the insult, suggesting that microglia may be an attractive target for pharmacologic intervention. The cerebrospinal fluid (CSF) of Alzheimer disease (AD) patients contains antibodies that recognize activated microglia in the developing rat and in the ischemic gerbil brain. These results suggest that AD shares common mechanisms related to the activation of microglia with both these experimental models. In vitro, the xanthine derivative propentofylline (PPF) depresses the production of reactive oxygen intermediates produced by macrophages. To appreciate in vivo interactions of PPF, two models were employed: developing rats and adult gerbils exposed to ischemia. Newborn rats were administered PPF (10 mg/kg) for 7 days. Gerbils were exposed to 5 min of transient forebrain ischemia and received PPF (10 mg/kg) 24 h later until the day before sacrifice. Animals were sacrificed at 7 or 14 days after reperfusion. Brains were processed for immunocytochemistry. Reactive microglia were visualized with monoclonal antibodies OX 18 and OX42 or AD-CSF microglia antibodies. In the case of ischemia, an antibody against the amyloid precursor protein (APP) (residues 676–695) was included. Newborn rats receiving PPF for 7 days displayed a dramatic reduction in the number of activated microglia compared with untreated littermates. Ischemic control in gerbils showed complete nerve death, accumulations of APP, and enhanced microglial reactivity. In gerbils receiving PPF, APP accumulation was absent or very slight, and activated microglia were downregu-lated. The ability of PPF to interfere with activated microglia suggests that this agent may be useful for slowing progressive nerve cell death associated with AD, which is considered to be largely influenced by pathologic glial reactions.

ISCHEMIC NEURONAL DAMAGE : HOW DOES MILD HYPOTHERMIA MODULATE IT ?

The significance of mild hypothermia as a therapeutic measure for ischemic brain damage is presented on the basis of different experimental results. An extracellular glutamate surge, a sustained activation of N-methyl-D-aspartate (NMDA) receptors, and an enhancement of DNA binding activity to transcription factor AP-1, all being key items directly linked to excitotoxic neuronal damage, are deeply affected by slightly lowering temperature (mild hypothermia [MH]). The cellular mechanism of MH seems rather nonspecific but tends to collectively involve these key items rendering neurons resistant to ischemic damage. Clinical application of MH should be a great challenge to relieve deadly effects on central neurons.

Cerebrospinal fluid microglial antibodies: Potential diagnostic markers for immune mechanisms in Alzheimer's disease

Hallmark lesions of Alzheimers disease (AD) are filled with reactive immunocompetent microglia, suggesting that immunological aberrations may participate in the pathophysiology of this disorder. Microglia may participate in the initial stages of neurodegeneration before the onset of dementia. If immune mediated processes are closely linked to neuronal breakdown it would be of importance to have a reliable means to detect these processes. Serum and cerebrospinal fluid (CSF) antibodies are discussed as such potential sources. The serendipitous use of the developing rat central nervous system (CNS) to screen CSF antibrain antibodies produced some unexpected findings. Firstly, CSF antibodies of AD and other dementia patients recognized distinctly different neuronal structures in the developing rat brain. Secondly, some AD CSF recognized fiber networks whereas others recognized amoeboid microglial cells. The same AD CSF which recognized amoeboid microglia cells also specifically marked activated microglia and neural macrophages in experimentally induced lesions. AD CSF microglial antibodies appear to be significant in view of the increasing association between microglia and neurogenerative processes in AD. In addition, CSF microglial antibodies are present in numerous at-risk descendants of familial AD patients. Some have subsequently developed the disorder. These findings together with the fact that microglial antibodies are usually found in the early stages of AD suggest that AD CSF microglial antibodies could be of value in detecting neurodegenerative processes before the onset of dementia. These findings add further support to the concept that inflammation and similar immune mechanisms may contribute to AD pathogenesis.

Adult-onset calorie restriction attenuates kainic acid excitotoxicity in the rat hippocampal slice.

Lifelong calorie restriction is the only known intervention that has been shown to consistently increase life span and reduce the effects of aging on the brain. Given the difficulties of replicating lifelong calorie restriction within human populations, we have sought to assess the effects of short-term adult-onset calorie restriction upon acute excitotoxic insults in the rat hippocampus. Adult animals (approximately 6 months of age) underwent calorie restriction (alternate day feeding) for 7-10 weeks. Utilizing both electrophysiological and immunocytochemical techniques, we report that calorie restriction had no effect upon long-term potentiation (LTP), a measure of neuronal function. In control animals, application of kainic acid (20 microM) resulted in only 35% recovery of CA1 population spikes post-insult. However calorie-restricted animals showed significantly improved recovery after kainic acid treatment (64%). This data was supported by immunocytochemical studies which noted widespread loss of microtubule-associated protein (MAP 2) immunoreactivity in control slices following treatment with kainic acid; however MAP 2 staining was preserved in the CA1 and CA3 regions of calorie-restricted animals. Interestingly there was no significant difference in the recovery of population spikes between groups when slices were treated with N-methyl-d-aspartate (15 microM). We conclude that short-term adult-onset calorie restriction does not alter normal neuronal function and serves to protect the hippocampus from acute kainic acid excitotoxicity.

Modulation of glial cell signaling by adenosine and pharmacological reinforcement. A neuroprotective strategy

In view of the increasing evidence that a pathological glial activation plays a significant role in the development of neurodegenerative diseases, we investigated the underlying molecular signaling as a possible target for a pharmacological therapy. Here, we are particularly focusing on the endogenous modulation of the Ca2+ and cyclic nucleotide-dependent signaling by the nucleoside adenosine and its reinforcement by the xanthine derivative propentofylline (PPF). As an experimental model, we used cultured rat microglial cells and astrocytes that are immature, show a high proliferation rate, and resemble in several aspects pathologically activated glial cells. A prolonged increase of the cellular cAMP level favored the differentiation of cultured astrocytes and associated properties required for the physiological nerve cell function. On the other hand, a strengthening of the cyclic nucleotide-dependent signaling inhibited potentially neurotoxic properties of cultured microglial cells. Similar effects were obtained by treatment with propentofylline, which mimicked modulatory adenosine effects and increased the intracellular level of cAMP and cGMP. Such a pharmacological glial cell conditioning, obtained by modifying the strength and the timing of these second messengers, may provide a therapy of neurodegenerative diseases in which a pathological activation of microglial cells and astrocytes is discussed to play a pathogenic role.

Sialic acid, homocysteine and CRP: potential markers for dementia.

To investigate whether sialic acid could discriminate between healthy age matched controls and patients with dementias of the Alzheimers type (AD), and pure vascular dementia (VaD). 27 patients and 51 controls were administered the Mini-Mental State Examination (MMSE) and had blood analyzed for levels of total sialic acid, total homocysteine (tHcy), and C-reactive protein (CRP). Significant differences were found between the mean MMSE scores for patients with dementia compared with controls. Sialic acid levels were significantly higher in patients with AD compared with controls and homocysteine levels were higher in VaD. Sialic acid levels discriminated between patients with dementia of the Alzheimers type and healthy controls only. The MMSE could discriminate between controls and patients with dementia but not between the subtypes and homocysteine was significant for patients with VaD.

Cerebrospinal fluid antimicroglial antibodies in Alzheimer disease: A putative marker of an ongoing inflammatory process

Immunocompetent microglia play an important role in the pathogenesis of Alzheimers disease (AD). Antimicroglial antibodies in the cerebrospinal fluid (CSF) in clinically diagnosed AD patients have been previously recorded. Here, we report the results of the analysis of the CSF from 38 autopsy cases: 7 with definite AD; 14 with mild and 10 with moderate Alzheimers type pathology; and 7 controls. Antimicroglial antibodies were identified in 70% of patients with definite AD, in 80% of patients with moderate and in 28% of patients with mild Alzheimers type pathology. CSF antimicroglial antibodies were not observed in any of the control cases. The results show that CSF antimicroglial antibodies are present in the majority of patients with definite AD and also in cases with moderate Alzheimers type changes. They may also indicate dysregulation of microglial function. Together with previous observations, these findings indicate that compromised immune defense mechanisms play an important role in the pathogenesis of AD.