Implications of high homocysteine levels in migraine pain: An experimental study of the excitability of peripheral meningeal afferents in rats with hyperhomocysteinemia.

OBJECTIVES: Investigation of chronic homocysteine action on the excitability and N-methyl-D-aspartate (NMDA) sensitivity of the peripheral trigeminovascular system of rats. BACKGROUND: Migraine is a neurological disease that affects 15%-20% of the general population. Epidemiological observations show that an increase of the sulfur-containing amino acid homocysteine in plasma-called hyperhomocysteinemia-is associated with a high risk of migraine, especially migraine with aura. In animal studies, rats with hyperhomocysteinemia demonstrated mechanical allodynia, photophobia, and anxiety, and higher sensitivity to cortical spreading depression. In addition, rats with hyperhomocysteinemia were more sensitive in a model of chronic migraine induced by nitroglycerin which indicated the involvement of peripheral nociceptive mechanisms. The present work aimed to analyze the excitability of meningeal afferents and neurons isolated from the trigeminal ganglion of rats with prenatal hyperhomocysteinemia. METHODS: Experiments were performed on male rats born from females fed with a methionine-rich diet before and during pregnancy. The activity of meningeal afferents was recorded extracellularly in hemiskull preparations ex vivo and action potentials were characterized using cluster analysis. The excitability of trigeminal ganglion neurons was assessed using whole-cell patch clamp recording techniques and calcium imaging studies. Meningeal mast cells were stained using toluidine blue. RESULTS: The baseline extracellular recorded electrical activity of the trigeminal nerve was higher in the hyperhomocysteinemia group with larger amplitude action potentials. Lower concentrations of KCl caused an increase in the frequency of action potentials of trigeminal afferents recorded in rat hemiskull ex vivo preparations. In trigeminal ganglion neurons of rats with hyperhomocysteinemia, the current required to elicit at least one action potential (rheobase) was lower, and more action potentials were induced in response to stimulus of 2 × rheobase. In controls, short-term application of homocysteine and its derivatives increased the frequency of action potentials of the trigeminal nerve and induced Ca2+ transients in neurons, which are associated with the activation of NMDA receptors. At the same time, in rats with hyperhomocysteinemia, we did not observe an increased response of the trigeminal nerve to NMDA. Similarly, the parameters of Ca2+ transients induced by NMDA, homocysteine, and its derivatives were not changed in rats with hyperhomocysteinemia. Acute incubation of the meninges in homocysteine and homocysteinic acid did not change the state of the mast cells, whereas in the model of hyperhomocysteinemia, an increased degranulation of mast cells in the meninges was observed. CONCLUSIONS: Our results demonstrated higher excitability of the trigeminal system of rats with hyperhomocysteinemia. Together with our previous finding about the lower threshold of generation of cortical spreading depression in rats with hyperhomocysteinemia, the present data provide evidence of homocysteine as a factor that increases the sensitivity of the peripheral migraine mechanisms, and the control of homocysteine level may be an important strategy for reducing the risk and/or severity of migraine headache attacks.

On the relationship between the properties of planar structures of non-ionic surfactants and their vesicular analogues - Niosomes.

In recent years, the study of niosomes as nanocarriers alternative to liposomes has received increasing attention. In contrast to well-studied liposome membranes, many aspects of the behavior of analogous niosome bilayers have not been studied. This paper considers one of these aspects related to the communication between the physicochemical properties of planar and vesicular objects. We present the first results of comparative studies of Langmuir monolayers of binary and ternary (with cholesterol) mixtures of non-ionic surfactants based on sorbitan esters and niosomal structures assembled from the same materials. The Thin-Film Hydration (TFH) method in the gentle shaking version was used to produce the particles of large sizes, while small unilamellar high quality vesicles with a unimodal distribution of particles were prepared by TFH using ultrasonic treatment and extrusion. An analysis of the structural organization and phase state of monolayers based on compression isotherms and supplemented by thermodynamic calculations, as well as the results of determining the particle morphology, polarity and microviscosity of niosome shells, made it possible to obtain fundamental data on the intermolecular interactions of the components and their packing in shells and to relate these data to the properties of niosomes. This relationship can be used to optimize the composition of niosome membranes and predict the behavior of these vesicular systems. It was shown that cholesterol excess creates regions of bilayers with increased rigidity (like "lipid rafts"), which hinders the process of folding film fragments into small niosomes.

Dual-Responsive and Reusable Optical Sensors Based on 2,3-Diaminoquinoxalines for Acidity Measurements in Low-pH Aqueous Solutions.

This work is focused on the age-old challenge of developing optical sensors for acidity measurements in low-pH aqueous solutions (pH < 5). We prepared halochromic (3-aminopropyl)amino-substituted quinoxalines QC1 and QC8 possessing different hydrophilic-lipophilic balance (HLB) and investigated them as molecular components of pH sensors. Embedding the hydrophilic quinoxaline QC1 into the agarose matrix by sol-gel process allows for fabrication of pH responsive polymers and paper test strips. The emissive films thus obtained can be used for a semi-quantitative dual-color visualization of pH in aqueous solution. Being exposed to acidic solutions with pH in the range of 1-5, they rapidly give different color changes when the analysis is performed in daylight or under irradiation at 365 nm. Compared with classical non-emissive pH indicators, these dual-responsive pH sensors allow for an increase in the accuracy of pH measurements, particularly in complex environmental samples. pH indicators for quantitative analysis can be prepared by the immobilization of amphiphilic quinoxaline QC8 using Langmuir-Blodgett (LB) and Langmuir-Schäfer (LS) techniques. Compound QC8 possessing two long alkyl chains (n-C8H17) forms stable Langmuir monolayers at the air-water interface, and these monolayers can be successfully transferred onto hydrophilic quartz and hydrophobic polyvinylchlorid (PVC) substrates using LB and LS techniques, respectively. The 30-layer films thus obtained are emissive, reveal excellent stability, and can be used as dual-responsive pH indicators for quantitative measurements in real-world samples with pH in the range of 1-3. The films can be regenerated by immersing them in basic aqueous solution (pH = 11) and can be reused at least five times.

Synthesis and Self-Assembly of ß-Octa[(4-Diethoxyphosphoryl)phenyl]porphyrins.

The ß-substituted porphyrinoids commonly used to form functional assembled systems in nature yet are still scarcely used in material chemistry probably due to the laborious synthesis of these compounds. In this work, ß-octa[(4-diethoxyphosphoryl)phenyl]porphyrin (2HOPPP) and its metal (Zn(II), Cd(II), Cu(II), and Ni(II)) complexes were prepared in good yields. These highly soluble chromophores were characterized in solution using spectroscopic (NMR, UV-vis, fluorescence), electrochemical, and spectroelectrochemical methods. Attachment of the electron-deficient residue (ArP(O)(OEt)2) to the porphyrin macrocycle leads to easier reductions and harder oxidations of the macrocycle for all complexes studied as compared to corresponding meso-tetra[4-(diethoxyphosphoryl)phenyl]porphyrin derivatives reported previously. We demonstrated that the strong electron-deficient character of the MOPPP porphyrins results principally from the increase in the number of electron-withdrawing groups at the periphery of the tetrapyrrolic macrocycle. Electron-deficient porphyrins are highly required in supramolecular and material chemistry in part due to their ability to form supramolecular assemblies via the coordination of axial ligands to the central metal atom. According to single-crystal X-ray data, ZnOPPP forms in the crystalline phase dimers in which each of the two tetrapyrrolic macrocycles is connected through an unusual combination of hydrogen bonding of two phosphoryl groups and the water molecules axially coordinated to the zinc atom of the partner molecule. The involvement of water molecules in porphyrin binding allows for an increase of distance between two porphyrin mean N4 planes, up to 4.478 Å. The offset of phosphoryl groups attached to the macrocycle through a 1,4-phenylene spacer withdraws the whole porphyrin macrocycle of one molecule from spatial overlap with the macrocycle of a partner molecule and increases the Zn-Zn distance up to 10.372 Å. This still unknown type of porphyrin dimers allows one to get deeper insights into the organization of naturally occurring tetrapyrrolic macrocycles. ZnOPPP also forms a labile dimeric complex in 5.3 × 10-7-5.8 × 10-5 M chloroform solutions. In contrast, other complexes prepared in this work exist as monomeric species under these experimental conditions. The self-association constant of ZnOPPP has been determined by electronic absorption spectroscopy.

Testing the Role of Glutamate NMDA Receptors in Peripheral Trigeminal Nociception Implicated in Migraine Pain.

The pro-nociceptive role of glutamate in the CNS in migraine pathophysiology is well established. Glutamate, released from trigeminal afferents, activates second order nociceptive neurons in the brainstem. However, the function of peripheral glutamate receptors in the trigeminovascular system suggested as the origin site for migraine pain, is less known. In the current project, we used calcium imaging and patch clamp recordings from trigeminal ganglion (TG) neurons, immunolabelling, CGRP assay and direct electrophysiological recordings from rat meningeal afferents to investigate the role of glutamate in trigeminal nociception. Glutamate, aspartate, and, to a lesser extent, NMDA under free-magnesium conditions, evoked calcium transients in a fraction of isolated TG neurons, indicating functional expression of NMDA receptors. The fraction of NMDA sensitive neurons was increased by the migraine mediator CGRP. NMDA also activated slowly desensitizing currents in 37% of TG neurons. However, neither glutamate nor NMDA changed the level of extracellular CGRP. TG neurons expressed both GluN2A and GluN2B subunits of NMDA receptors. In addition, after removal of magnesium, NMDA activated persistent spiking activity in a fraction of trigeminal nerve fibers in meninges. Thus, glutamate activates NMDA receptors in somas of TG neurons and their meningeal nerve terminals in magnesium-dependent manner. These findings suggest that peripherally released glutamate can promote excitation of meningeal afferents implicated in generation of migraine pain in conditions of inherited or acquired reduced magnesium blockage of NMDA channels and support the usage of magnesium supplements in migraine.

Alcohol metabolite acetic acid activates BK channels in a pH-dependent manner and decreases calcium oscillations and exocytosis of secretory granules in rat pituitary GH3 cells.

Acetaldehyde and acetic acid/acetate, the active metabolites of alcohol (ethanol, EtOH), generate actions of their own ranging from behavioral, physiological, to pathological/cancerogenic effects. EtOH and acetaldehyde have been studied to some depth, whereas the effects of acetic acid have been less well explored. In this study, we investigated the effect of acetic acid on big conductance calcium-activated potassium (BK) channels present in GH3 rat pituitary tumor cells in more detail. In whole cell voltage clamp recordings, extracellular application of acetic acid increased total outward currents in a dose-dependent manner. This effect was prevented after the application of the specific BK channel blocker paxilline. Acetic acid action was pH-dependent-in whole cell current and single BK channel recordings, open probability (Po) was significantly increased by extracellular pH reduction and decreased by neutral or base pH. Acetic acid hyperpolarized the membrane potential, whereas acidic physiological solution had a depolarizing effect. Moreover, acetic acid reduced calcium (Ca2+) oscillations and exocytosis of growth hormone contained secretory granules from GH3 cells. These effects were partially prevented by BK inhibitors-tetraethylammonium or paxillin. In conclusion, our experiments indicate that acetic acid activates BK channels in GH3 cells which eventually contribute to acetic acid-induced membrane hyperpolarization, cessation of Ca2+ oscillations, and decrease of growth hormone release.

Protective Effects of Hydrogen Sulfide Against the ATP-Induced Meningeal Nociception.

We previously showed that extracellular ATP and hydrogen sulfide (H2S), a recently discovered gasotransmitter, are both triggering the nociceptive firing in trigeminal nociceptors implicated in migraine pain. ATP contributes to meningeal nociception by activating the P2X3 subunit-containing receptors whereas H2S operates mainly via TRP receptors. However, H2S was also proposed as a neuroprotective and anti-nociceptive agent. This study aimed to test the effect of H2S on ATP-mediated nociceptive responses in rat meningeal afferents and trigeminal neurons and on ATP-induced degranulation of dural mast cells. Electrophysiological recording of trigeminal nerve activity in meninges was supplemented by patch-clamp and calcium imaging studies of isolated trigeminal neurons. The H2S donor NaHS induced a mild activation of afferents and fully suppressed the subsequent ATP-induced firing of meningeal trigeminal nerve fibers. This anti-nociceptive effect of H2S was specific as an even stronger effect of capsaicin did not abolish the action of ATP. In isolated trigeminal neurons, NaHS decreased the inward currents and calcium transients evoked by activation of ATP-gated P2X3 receptors. Moreover, NaHS prevented ATP-induced P2X7 receptor-mediated degranulation of meningeal mast cells which emerged as triggers of migraine pain. Finally, NaHS decreased the concentration of extracellular ATP in the meningeal preparation. Thus, H2S exerted the multiple protective actions against the nociceptive effects of ATP. These data highlight the novel pathways to reduce purinergic mechanisms of migraine with pharmacological donors or by stimulation production of endogenous H2S.

Interfacial self-assembly of functional bilayer templates comprising porphyrin arrays and graphene oxide.

Fabricating of solid-supported hybrid nanostructures remains a challenging problem because it is difficult to control all interfacial interactions influencing the structure and stability of these systems. The most widely used approach to solving this problem is a bottom-up assembly on the surface templates such as self-assembled monolayers (SAMs). Herein we suggest an alternative approach to tailoring solid surfaces by a formation of an interlayer anchoring the nanostructured film to the solid substrate. We formed a multifunctional bilayer template (MBT), comprising an adhesive monolayer of graphene oxide and a functional ordered monolayer of metal organic compound (Zinc-tetra(4-pyridyl)porphyrin) directing further bottom-up growth of the nanostructures. The one-step assembly of MBT proceeded spontaneously at the air/water interface and was monitored by an in-situ fiber optic absorption and fluorescence spectroscopy in a Langmuir trough. Dilatation surface rheology was applied to study the evolution of molecular organization of the monolayers upon adding the zinc ions, GO and their mixture into the subphase. The MBT templates were used for the assembly of porphyrin-based SURMOFs with two different structures. Our strategy makes it possible to assemble surface-anchored nanostructures avoiding the use of SAMs and it can be extended to other types of ultrathin hybrid systems.

A metal-responsive interdigitated bilayer for selective quantification of mercury(ii) traces by surface plasmon resonance.

Reusable surface plasmon resonance chips allowing the quantitative and selective detection of mercury(ii) ions in water at the 0.01 nM level are reported. The surface-modified gold sensor consists of a rarefied self-assembled monolayer of octanethiol topped with a Langmuir-Blodgett monolayer of an amphiphilic and highly-specific chelator. The interdigitated architecture confers to the bilayer a high packing density, surface coverage, and binding-group accessibility.

Colorimetric Hg2+ sensing in water: from molecules toward low-cost solid devices.

A new colorimetric molecular sensor allowing for cheap, fast, sensitive, and highly selective naked-eye detection of Hg(2+) in water is described. This molecule combines a 1,8-diaminoanthraquinone signaling subunit and phosphonic acid esters that confer the water solubility to the dye (R = H). A ready-to-use colorimetric solid sensor was obtained by incorporating an amphiphilic analog (R = OC(12)H(25)) exhibiting similar binding properties and optical responses in an agarose film.