Maryellen F. Eckenhoff

Inhaled Anesthetic Enhancement of Amyloid-β Oligomerization and Cytotoxicity

Background: The majority of surgical patients receive inhaled anesthetics, principally small haloalkanes and haloethers. Long-term cognitive problems occur in the elderly subsequent to anesthesia and surgery, and previous surgery might also be a risk factor for neurodegenerative disorders like Alzheimer and Parkinson disease. The authors hypothesize that inhaled anesthetics contribute to these effects through a durable enhancement of peptide oligomerization. Methods: Light scattering, filtration assays, electron microscopy, fluorescence spectroscopy and size-exclusion chromatography was used to characterize the concentration-dependent effects of halothane, isoflurane, propofol, and ethanol on amyloid &bgr; peptide oligomerization. Pheochromocytoma cells were used to characterize cytotoxicity of amyloid oligomers with and without the above anesthetics. Results: Halothane and isoflurane enhanced amyloid beta oligomerization rates and pheochromocytoma cytotoxicity in vitro through a preference for binding small oligomeric species. Ethanol and propofol inhibited oligomerization at low concentration but enhanced modestly at very high concentration. Neither ethanol nor propofol enhanced amyloid &bgr; toxicity in pheochromocytoma cells. Conclusions: Inhaled anesthetics enhance oligomerization and cytotoxicity of Alzheimer disease-associated peptides. In addition to the possibility of a general mechanism for anesthetic neurotoxicity, these results call for further evaluation of the interaction between neurodegenerative disorders, dementia, and inhalational anesthesia.

Brain and behavior changes in 12-month-old Tg2576 and nontransgenic mice exposed to anesthetics

Inhaled anesthetics have been shown to increase the aggregation of amyloid beta in vitro through the stabilization of intermediate toxic oligomers, which are thought to contribute to neurocognitive dysfunction in Alzheimers disease. Inhaled anesthetics may escalate cognitive dysfunction through enhancement of these intermediate oligomer concentrations. We intermittently exposed 12-month-old Tg2576 transgenic mice and nontransgenic littermates to isoflurane and halothane for 5 days. Cognitive function was measured before and after anesthetic exposures using the Morris Water Maze; amyloid beta plaque burden and caspase-3 mediated apoptosis were quantified by immunohistochemistry. At 12 months of age, anesthetic exposure did not further enhance cognitive decline in the transgenic mice. Immunohistochemistry, however, revealed that the halothane-exposed Tg2576 mice had more amyloidopathy than the isoflurane treated mice or the nonexposed transgenic mice. Isoflurane exposure impaired cognitive function in the nontransgenic mice, implying an alternative pathway for neurodegeneration. These findings indicate that inhaled anesthetics influence cognition and amyloidogenesis, but that the mechanistic relationship remains unclear.

Human Alzheimer and inflammation biomarkers after anesthesia and surgery

Background: The prevalence of postoperative cognitive disturbance, coupled with growing in vitro, cell, and animal evidence suggesting anesthetic effects on neurodegeneration, calls for additional study of the interaction between surgical care and Alzheimer neuropathology. The authors studied human cerebrospinal fluid (CSF) biomarkers during surgery. Methods: Eleven patients undergoing idiopathic nasal CSF leak correction were admitted to this Institutional Review Board-approved study. Lumbar subarachnoid catheters were placed before the procedure. Anesthesia was total intravenous propofol or remifentanil or inhalational sevoflurane, depending on provider choice. CSF samples were taken after catheter placement (base), at procedure end (0 h), and then at 6, 24, and 48 h. CSF was analyzed using xMAP Luminex immunoassay (Luminex, Austin, TX). Results: Of the 11 patients (age range, 53 ± 6 yr), 8 were women; 4 received intravenous anesthesia, 6 sevoflurane, and 1 mixed. Procedures lasted 6.4 ± 2 h. Mean CSF amyloid-&bgr;(1–42) remained unchanged, but total-tau and phosphorylated-tau181P increased progressively until at least 48 h. Total-tau, phosphorylated-tau, or amyloid-&bgr;(1–42) concentrations were not different between anesthetic groups. CSF interleukin-10, S100Beta, and tumor necrosis factor &agr; were increased similarly in both anesthetic groups at 24 h, but interleukin-6 was increased more in the inhalational group. Conclusion: These data indicate a robust neuroinflammatory response, including not only the usual markers (interleukin-6, tumor necrosis factor &agr;, interleukin-10), but also S100Beta and tau, markers of injury. The total-tau/amyloid-&bgr;(1–42) ratio increased in a pattern consistent with Alzheimer disease, largely because of an increase in total-tau rather than a decline in amyloid-&bgr;(1–42). The differences in CSF interleukin-6 concentrations suggest that anesthetic management may make a difference in neuroinflammatory response.

Consensus statement: First International Workshop on Anesthetics and Alzheimer's disease.

In order to review the current status of the potential relationship between anesthesia and Alzheimer’s disease, a group of scientists recently met in Philadelphia for a full day of presentations and discussions. This special article represents a consensus view on the possible link between Alzheimer’s disease and anesthesia and the steps required to test this more definitively.

Dantrolene ameliorates cognitive decline and neuropathology in Alzheimer triple transgenic mice.

Disruption of intracellular calcium homeostasis via abnormal and excessive activation of ryanodine receptors plays an important role in the neuropathology of Alzheimers disease. We investigated the therapeutic effect of dantrolene, a ryanodine receptor antagonist, on cognitive dysfunction and neuropathology in the triple transgenic Alzheimer mouse model (3xTg-AD). 3xTg-AD mice were treated with dantrolene from 2 to 13 months of age. Learning and memory were measured with the Morris Water Maze at 6, 10, and 13 months of age. Amyloid and tau neuropathology and biomarkers for synaptic dysfunction and neurodegeneration were examined in the brain using immunoblotting or immunohistochemistry. Dantrolene treatment for 11 months significantly reduced both memory deficits and amyloid plaque load in the hippocampus in 13-month-old 3xTg-AD mice. Dantrolene treatment, however, had no effect on phosphorylated tau, phosphorylated or total GSK-3β, synaptic markers, or mitochondrial or cytosolic cytochrome C. Our results suggest that dantrolene significantly improves cognition in a murine model of Alzheimers disease and is associated with a reduction in amyloid plaque burden, forming the basis for a novel therapeutic approach for Alzheimers disease.

Anesthetic-Induced Neurodegeneration Mediated via Inositol 1,4,5-Trisphosphate Receptors

The commonly used general anesthetic isoflurane induces widespread neurodegeneration in the developing mammalian brain through poorly understood mechanisms. We have investigated whether excessive Ca2+ release from the endoplasmic reticulum via overactivation of inositol 1,4,5-trisphosphate receptors (InsP3Rs) is a contributing factor in such neurodegeneration in rodent primary cultured neurons and developing rat brain. We also investigated the correlation between isoflurane exposure and cognitive decline in rats at 1 month of age. Our results show that isoflurane increases cytosolic calcium in the primary cortical neurons through release from the endoplasmic reticulum and influx from the extracellular space. Pharmacological inhibition of InsP3R activity and knockdown of its expression nearly abolishes the isoflurane-mediated elevation of the cytosolic calcium concentration and cell death in rodent primary cortical and hippocampal neurons. Inhibition of InsP3R activity by its antagonist xestospongin C significantly inhibits neurodegeneration induced by isoflurane at clinically used concentration in the developing brain of postnatal day 7 rats. Moreover, our results show that isoflurane activates β-site amyloid β precursor protein-cleaving enzyme via activation of the InsP3R. We also noted that mice exposed to isoflurane during early postnatal development showed transient memory and learning impairments, which did not correlate well with the noted neuropathological defects. Taken together, our results suggest that Ca2+ dysregulation through overactivation of the InsP3R may be a contributing factor in the mechanism of isoflurane-induced neurodegeneration in rodent neuronal cell culture and during brain development.

Anesthesia and the Old Brain

The perioperative period may have long-term consequences on cognitive function in the elderly patient. In this special article, we summarize the rationale and evidence that the anesthetic per se is a contributor. The evidence at this point is considered suggestive and further research is needed, especially in humans.

A Presenilin-1 Mutation Renders Neurons Vulnerable to Isoflurane Toxicity

BACKGROUND:Isoflurane, a commonly used inhaled anesthetic, induces apoptosis in rat pheochromocytoma neurosecretory cells (PC12) in a concentration- and time-dependent manner via an as yet unknown mechanism. We hypothesize that isoflurane induces apoptosis by causing abnormal calcium release from the endoplasmic reticulum (ER) via activation of inositol 1,4,5-trisphosphate (IP3) receptors. A presenilin-1 (PS1) mutation associated with familial Alzheimer’s disease was shown to increase the activity of IP3 receptors, and therefore may render cells vulnerable to isoflurane-induced cytotoxicity. Sevoflurane and desflurane have less ability to disrupt intracellular calcium homeostasis; and thus we predict they will cause less cytotoxicity. METHODS:PC12 cells transfected with wild type, vector alone (Vector) or mutated PS1 (L286V) were treated with equivalent of 1 MAC of isoflurane, sevoflurane, and desflurane for 12 h. Mitochondria redox activity (MTT reduction) and lactate dehydrogenase release assays were performed to evaluate cell viability. Changes of calcium concentration in cytosolic space ([Ca2+]c) and production of reactive oxygen species (ROS) were determined after exposing different types of cells to various inhaled anesthetics. We also determined the effects of IP3 receptor antagonist xestospongin C on isoflurane-induced cytotoxicity and calcium release from the ER in L286V PC12 cells, and in rat primary cortical neurons. RESULTS:Isoflurane at 1 MAC for 12 h induced cytotoxicity in L286V but not wild type or vector PC12 cells, and also caused greater and faster increase of peak [Ca2+]c in the L286V cells. Xestospongin C significantly attenuated isoflurane cytotoxicity in both L286V cells and primary cortical neurons and inhibited the calcium release from the ER in L286V cells. Isoflurane did not induce significant changes of ROS production in any type of PC12 cells. Sevoflurane and desflurane at equivalent exposure to isoflurane did not induce similar cytotoxicity or increase of peak [Ca2+]c in L286V PC12 cells. CONCLUSION:Our results show that the L286V PS1 mutation augments the isoflurane-induced [Ca2+]c increase via calcium release from intracellular stores which, in turn, renders the cells vulnerable to isoflurane neurotoxicity. ROS production was not involved in isoflurane-induced neurotoxicity. Sevoflurane and desflurane, at equivalent exposure to isoflurane, did not induce a similar increase of [Ca2+]c or neurotoxicity in L286V PC12 cells.

Differential General Anesthetic Effects on Microglial Cytokine Expression

Post-operative cognitive dysfunction has been widely observed, especially in older patients. An association of post-operative cognitive dysfunction with the neurodegenerative diseases, such as Alzheimers disease, has been suggested. Neuroinflammation contributes to Alzheimer pathology, through elevated pro-inflammatory cytokines and microglial activation in the CNS leading to neuronal damage, synaptic disruption and ultimately cognitive dysfunction. We compare the effects of three different, clinically-used, anesthetics on microglial activation with, and without, the prototypical inflammatory trigger, lipopolysaccharide (LPS). Microglial BV-2 cell cultures were first exposed to isoflurane, sevoflurane (each at 2 concentrations) or propofol for 6 h, and cytokine levels measured in lysates and media. The same experiments were repeated after 1 h LPS pre-treatment. We found; 1) anesthetics alone have either no or only a small effect on cytokine expression; 2) LPS provoked a large increase in microglia cytokine expression; 3) the inhaled anesthetics either had no effect on LPS-evoked responses or enhanced it; 4) propofol nearly eliminated the LPS pro-inflammatory cytokine response and improved cell survival as reflected by lactate dehydrogenase release. These data suggest that propofol may be a preferred anesthetic when it is desirable to minimize neuroinflammation.

Acute Anemia Elicits Cognitive Dysfunction and Evidence of Cerebral Cellular Hypoxia in Older Rats with Systemic Hypertension

Background:Postoperative cognitive dysfunction occurs frequently after cardiac, major vascular, and major orthopedic surgery. Aging and hypertensive cerebrovascular disease are leading risk factors for this disorder. Acute anemia, common to major surgery, has been identified as a possible contributor to postoperative cognitive dysfunction. The effect of hypoxia upon cognition and the cellular and molecular processes involved in learning and memory has been well described. Cerebrovascular changes related to chronic hypertension may expose cells to increased hypoxia with anemia. Methods:Young to aged spontaneously hypertensive rats underwent testing for visuospatial memory and learning in the Morris water maze, measurement of cerebral tissue oxygenation via tissue oxygen probe, and measurement of hypoxia-sensitive genes and proteins, under conditions of sham and experimental isovolemic anemia. Results:Acute isovolemic anemia elicited evidence of aging-dependent visuospatial working memory and learning impairment. Isovolemic anemia did not result in cerebral tissue hypoxia, when measured via tissue oxygen probe. Evidence of cellular hypoxia was, however, identified in response to the anemic challenge, as hypoxia-sensitive genes and proteins were up-regulated. Importantly, cellular hypoxic gene responses were increased with anemia in an age-dependent manner in this model of aging with chronic hypertension. Conclusions:In a translational model of chronic hypertension, clinically relevant levels of acute anemia were associated with an age-dependent visuospatial working memory and learning impairment that was matched by an age-dependent cellular sensitivity to anemic hypoxia. These data offer support for a possible link between anemic hypoxia and postoperative cognitive dysfunction in humans.