Dietrich E. Lorke

Methylene blue and Alzheimer's disease.

The relationship between methylene blue (MB) and Alzheimers disease (AD) has recently attracted increasing scientific attention since it has been suggested that MB may slow down the progression of this disease. In fact, MB, in addition to its well characterized inhibitory actions on the cGMP pathway, affects numerous cellular and molecular events closely related to the progression of AD. Currently, MB has been shown to attenuate the formations of amyloid plaques and neurofibrillary tangles, and to partially repair impairments in mitochondrial function and cellular metabolism. Furthermore, various neurotransmitter systems (cholinergic, serotonergic and glutamatergic), believed to play important roles in the pathogenesis of AD and other cognitive disorders, are also influenced by MB. Recent studies suggest that the combination of diverse actions of MB on these cellular functions is likely to mediate potential beneficial effects of MB. This has lead to attempts to develop novel MB-based treatment modalities for AD. In this review article, actions of MB on neurotransmitter systems and multiple cellular and molecular targets are summarized with regard to their relevance to AD.

Cellular and molecular actions of Methylene Blue in the nervous system.

Methylene Blue (MB), following its introduction to biology in the 19th century by Ehrlich, has found uses in various areas of medicine and biology. At present, MB is the first line of treatment in methemoglobinemias, is used frequently in the treatment of ifosfamide‐induced encephalopathy, and is routinely employed as a diagnostic tool in surgical procedures. Furthermore, recent studies suggest that MB has beneficial effects in Alzheimers disease and memory improvement. Although the modulation of the cGMP pathway is considered the most significant effect of MB, mediating its pharmacological actions, recent studies indicate that it has multiple cellular and molecular targets. In the majority of cases, biological effects and clinical applications of MB are dictated by its unique physicochemical properties including its planar structure, redox chemistry, ionic charges, and light spectrum characteristics. In this review article, these physicochemical features and the actions of MB on multiple cellular and molecular targets are discussed with regard to their relevance to the nervous system.

Entry of Oximes into the Brain: A Review

The passage of hydrophilic drugs, such as oxime acetylcholinesterase reactivators, into the central nervous system is restricted by the blood-brain and the blood-cerebrospinal fluid barriers. The present review summarizes morphological and functional properties of the blood-brain barrier, blood-cerebrospinal fluid barrier and cerebrospinal fluid-brain interface and reviews the existing data on brain entry of oximes. Due to the virtual absence of transcytosis, lack of fenestrations and unique properties of tight junctions in brain endothelial cells, the blood-brain barrier only allows free diffusion of small lipophilic molecules. Various carriers transport hydrophilic compounds and extrude potentially toxic xenobiotics. The blood-cerebrospinal fluid barrier is formed by the choroid plexus epithelium, whose tight junctions are more permeable than those of brain endothelial cells. The major function of plexus epithelium cells is active transport of ions for the production of the cerebrospinal fluid. The cerebrospinal fluid-brain interface is not a biological barrier and allows free diffusion. However, in contrast to passage via the blood-brain barrier or the blood-cerebrospinal fluid barrier, direct penetration from the cerebrospinal fluid into the brain is very slow, since much longer distances have to be covered. A bulk flow of brain interstitial fluid and cerebrospinal fluid speeds up exchange between these two fluid compartments. Oximes, by reactivating acetylcholinesterase, are important adjunct therapeutics in organophosphate poisoning. They are very hydrophilic and therefore cannot diffuse freely into the central nervous system. Changes in brain acetylcholinesterase activity, oxime concentration and some biological effects elicited by oxime administration in the periphery indicate, however, that oximes can gain access to the brain to a certain degree, probably by carrier-mediated transport, reaching in the brain about 4-10% of their respective plasma levels. The clinical relevance of this effect is hotly debated. Possible strategies to improve brain penetration of oximes are discussed.

Proliferation and apoptosis in the developing human neocortex

The cell kinetics of the developing central nervous system (CNS) is determined by both proliferation and apoptosis. In the human neocortex at week 6 of gestation, proliferation is confined to the ventricular zone, where mitotic figures and nuclear immunoreactivity for proliferating cell nuclear antigen (PCNA) are detectable. Cell division is symmetric, with both daughter cells reentering mitosis. At week 7, the subventricular zone, a secondary proliferative zone, appears. It mainly gives rise to local circuit neurons and glial cells. Around week 12, the ventricular and subventricular zones are thickest, and the nuclear PCNA label is strongest, indicating that proliferation peaks at this stage. Thereafter, asymmetric division becomes the predominant mode of proliferation, with one daughter cell reentering mitosis and the other one migrating out. Towards late gestation, the ventricular and subventricular zones almost completely disappear and proliferation shifts towards the intermediate and subplate zones, where mainly glial cells are generated. A remnant of the subventricular zone with proliferative activity persists into adulthood. In general, proliferation follows a latero‐medial gradient in the neocortex lasting longer in its lateral parts. Apoptotic nuclei have been detected around week 5, occurring in low numbers in the ventricular zone at this stage. Apoptotic cell death increases around midgestation and then spreads throughout all cortical layers, with most dying cells located in the ventricular and subventricular zones. This spatial distribution of apoptosis extends into late gestation. During the early postnatal period, most apoptotic cells are still located in the subcortical layers. During early embryonic development, proliferation and apoptosis are closely related, and are probably regulated by common regulators. In the late fetal and early postnatal periods, when proliferation has considerably declined in all cortical layers, apoptosis may occur in neurons whose sprouting axons do not find their targets. Anat Rec 267:261–276, 2002.

Expression of the Na-K-2Cl-cotransporter NKCC1 during mouse development

Here we describe the expression pattern of the Na-K-2Cl-cotransporter NKKC1 during embryonal and early postnatal mouse development. During early stages hybridization signals were detected over single cells of the developing neuroepithelia, whereas the neuroepithelium of the basal telencephalon was labeled continuously. With ongoing differentiation a distinct pattern of hybridization became apparent, which switched from a neuronal to a more glial pattern in the adult. Outside the nervous system NKCC1 transcripts were present in many organs and were mostly confined to epithelia.

Differential expression of the estrogen receptor-related receptor γ in the mouse brain

To elucidate estrogen functions, the expression of the estrogen receptor-related receptor ERRγ, a novel orphan nuclear receptor regulating transcription via estrogen responsive elements, has been localized by in situ hybridization in adult murine brain. ERRγ transcripts were abundantly present in the isocortex, the olfactory system, cranial nerve nuclei and major parts of the coordination centers, e.g. reticular formation and major parts of the extrapyramidal motor systems. In addition, ERRγ expression was detected in trigeminal ganglion neurons. ERRγ distribution was clearly distinguished from that described for ERRα, for ERRβ, and for estrogen receptors (ER) pointing at functional differences between ERRγ and these receptors.

CT-guided blocks and neuroablation of the ganglion impar (Walther) in perineal pain: anatomy, technique, safety, and efficacy.

ObjectiveAn alternate approach to the ganglion impar was chosen to minimize the risk of adverse events. Efficacy of the procedure was evaluated. MethodsCharts and computed tomography (CT)-scans of patients who underwent block and neuroablation of the ganglion impar (Walther) between 2003 and 2007 were systematically reviewed with respect to adverse events and efficacy by rating pain intensity. A total of 76 blocks were performed, 48 of them being diagnostic blocks and 28 neuroablations. Chemical destruction was performed with ethanol, if pain recurred despite injection of local anesthetic. ResultsInterventional pain therapy was performed in 43 patients (age: 64.6±12.4 y, median 49.5 y, range: 36 to 86 y, male/female: 27/16) presenting with perineal pain of unknown origin (n=15), carcinoma of the prostate (n=8), colorectal carcinoma (n=7), postsurgery of thrombosis of perineal veins (n=3), postherpetic neuralgia (n=4), malformation of the spinal cord (n=2), vaginal protrusion (n=2), failed back surgery syndrome (n=1), and ablation of testis (n=1). CT-guided puncture was not associated with any adverse events and resulted in a reduction of numeric rating scale values from 8.2±1.6 to 2.2±1.6 (P<0.0001, 95% confidence interval 0.5) immediately at discharge and to 2.2±1.4 (P<0.0001, 95% confidence interval 0.4) at 4 months on follow up. DiscussionCT-guided block and neuroablation of the ganglion impar (Walther) results in a significant reduction of pain scores and carries virtually no hazards.

In vitro oxime protection of human red blood cell acetylcholinesterase inhibited by diisopropyl-fluorophosphate.

Oximes are enzyme reactivators used in treating poisoning with organophosphorus cholinesterase (AChE) inhibitors. The oxime dose which can be safely administered is limited by the intrinsic toxicity of the substances such as their own AChE‐inhibiting tendency. Clinical experience with the available oximes is disappointing. To meet this need, new AChE reactivators of potential clinical utility have been developed. The purpose of the study was to estimate in vitro both the intrinsic toxicity and the extent of possible protection conferred by established (pralidoxime, obidoxime, HI‐6, methoxime, trimedoxime) and experimental (K‐type) oximes, using diisopropyl‐fluoro‐phosphate (DFP) as an AChE inhibitor. The IC50 of DFP against human red blood cell AChE was determined (∼120 nm). Measurements were then repeated in the presence of increasing oxime concentrations, leading to an apparent increase in DFP IC50. Calculated IC50 values were plotted against oxime concentrations to obtain an IC50 shift curve. The slope of this shift curve (tanα) was used to quantify the magnitude of the protective effect (nm IC50 increase per µm oxime). We show that, in the case of a linear relationship between oxime concentration and IC50, the binding constant K, determined using the Schild equation, equals IC50/DFP/tanα. Based on the values of tanα and of the binding constant K, some of the new K‐oxime reactivators are far superior to pralidoxime (tanα = 0.8), obidoxime (1.5), HI‐6 (0.8), trimedoxime (2.9) and methoxime (5.9), with K‐107 (17), K‐108 (20), and K‐113 (16) being the outstanding compounds. Copyright

Eight new bispyridinium oximes in comparison with the conventional oximes pralidoxime and obidoxime: in vivo efficacy to protect from diisopropylfluorophosphate toxicity

In search for more efficacious reactivators of acetylcholinesterase (AChE) inhibited by organophosphorus compounds, experimental K‐oximes have been synthesized which show good in vitro efficacy. However, AChE inhibition by oximes themselves (as quantified by their intrinsic IC50) is the major cause of oxime toxicity and the dose‐limiting factor. To assess K‐oxime efficacy in vivo, the extent of protection from mortality induced by diisopropylfluorophosphate (DFP) was quantified by Cox survival analysis and compared with that of the clinically available oximes. Oximes were administered in an equitoxic dosage, i.e. half the LD01. Best protection was conferred by K‐27, reducing the relative risk of death (RR) to 16% of control RR (P ≤ 0.05), which was statistically significantly better (P ≤ 0.05) than all other tested oximes, except obidoxime, K‐53 and K‐75. The efficacy of obidoxime (RR = 0.19), K‐48 (RR = 0.28), K‐53 (RR = 0.22), K‐74 (RR = 0.38) and K‐75 (RR = 0.29) was significantly (P ≤ 0.05) better than that of 2‐PAM (RR = 0.62) and K‐113 (RR = 0.73). No significant protective effect was observed for K‐107 and K‐108. Our LD50 data show that K‐107, K‐108 and K‐113 (which strongly inhibit AChE in vitro) are in vivo markedly more toxic than all other oximes tested and can therefore only be safely administered at a low dosage which is insufficient to protect from DFP‐induced mortality. Dosage calculations based on in vitro IC50 measurements may therefore in future replace in vivo LD50 determinations, thereby reducing the number of animals required. Copyright

On the Interaction of β-Amyloid Peptides and α7-Nicotinic Acetylcholine Receptors in Alzheimer’s Disease

Deterioration of the cortical cholinergic system is a leading neurochemical feature of Alzheimers Disease (AD). This review summarizes evidence that the homomeric α7- nicotinic acetylcholine receptor (nAChR) plays a crucial role in the pathogenesis of this disease, which is characterized by amyloid-β (Aβ) accumulations and neurofibrillary tangles originating from of hyperphosphorylated tau protein. Aβ binds to α7-nAChRs with a high affinity, either activating or inhibiting this receptor in a concentration-dependent manner. There is strong evidence that α7-nAChRs are neuroprotective, reducing Aβ-induced toxicity; but co-localization of α7- nAChRs, Aβ and amyloid plaques also points to neurodegenerative actions. Aβ induces tau phosphorylation via α7-nAChR activation. Aβ influences hippocampus-dependent memory and long-term potentiation in a dose-dependent way: there is evidence that enhancement by picomolar Aβ concentrations is mediated by α7-nAChRs, whereas inhibition by nanomolar concentrations is independent of nAChRs and probably mediated by small Aβ42 oligomers. α7-nAChRs located on vascular smooth muscle cells and astrocytes are also involved in the pathogenesis of AD. Although these data strongly point to an important role of α7-nAChRs in the development of AD, dose-dependence of the effects, rapid desensitization of the receptor and dependence of the effects on Aβ aggregation (monomers, oligomers, fibrils) make it difficult to develop simple therapeutic strategies acting upon this receptor.