Assessment of the Potential Role of Muscle Spindle Mechanoreceptor Afferents in Chronic Muscle Pain in the Rat Masseter Muscle

Background

The phenotype of large diameter sensory afferent neurons changes in several models of neuropathic pain. We asked if similar changes also occur in “functional” pain syndromes.

Methodology/Principal Findings

Acidic saline (AS, pH 4.0) injections into the masseter muscle were used to induce persistent myalgia. Controls received saline at pH 7.2. Nocifensive responses of Experimental rats to applications of Von Frey Filaments to the masseters were above control levels 1–38 days post-injection. This effect was bilateral. Expression of c-Fos in the Trigeminal Mesencephalic Nucleus (NVmes), which contains the somata of masseter muscle spindle afferents (MSA), was above baseline levels 1 and 4 days after AS. The resting membrane potentials of neurons exposed to AS (n = 167) were hyperpolarized when compared to their control counterparts (n = 141), as were their thresholds for firing, high frequency membrane oscillations (HFMO), bursting, inward and outward rectification. The amplitude of HFMO was increased and spontaneous ectopic firing occurred in 10% of acid-exposed neurons, but never in Controls. These changes appeared within the same time frame as the observed nocifensive behaviour. Ectopic action potentials can travel centrally, but also antidromically to the peripheral terminals of MSA where they could cause neurotransmitter release and activation of adjacent fibre terminals. Using immunohistochemistry, we confirmed that annulospiral endings of masseter MSA express the glutamate vesicular transporter VGLUT1, indicating that they can release glutamate. Many capsules also contained fine fibers that were labelled by markers associated with nociceptors (calcitonin gene-related peptide, Substance P, P2X3 receptors and TRPV1 receptors) and that expressed the metabotropic glutamate receptor, mGluR5. Antagonists of glutamatergic receptors given together with the 2^nd injection of AS prevented the hypersensitivity observed bilaterally but were ineffective if given contralaterally.

Conclusions/Significance

Low pH leads to changes in several electrical properties of MSA, including initiation of ectopic action potentials which could propagate centrally but could also invade the peripheral endings causing glutamate release and activation of nearby nociceptors within the spindle capsule. This peripheral drive could contribute both to the transition to, and maintenance of, persistent muscle pain as seen in some “functional” pain syndromes.     

Cloning,high level expression and immunogenicity of 1163-1256 residues of C-terminal heavy chain of C. botulinum neurotoxin type E

Botulinum neurotoxins (BoNTs) inhibit neurotransmitter release from peripheral cholinergic synapses. BoNTs consist of a toxifying light chain and a heavy chain (HC) linked through a disulfide bond. In the present study we explored the immunogenicity and protective capability of the most effective part corresponding to 1163-1256 residues of botulinum type E neurotoxin HC gene. DNA encoding the 93 C-terminal amino acid of HC residues was synthesized with optimal codon usage for expression. These DNA fragments were ligated into a pLivSelect vector and subcloned into expression vector pET32a. Recombinant plasmids were then transformed into Escherichia coli strain BL21 DE3. The recombinant protein was purified by nickel affinity gel column chromatography. The HC1163-1256 was identified by antibodies raised against BoNT/E. HC1163-1256 was shown to bind with synaptotagmin and gangliosides, indicating that the expressed and purified HC1163-1256 protein retains a functionally active conformation. The immunization with recombinant protein induced a protection level in mice. The immunization with 2 μg of the recombinant protein induced a significant protection level in mice. In conclusion, availability of the recombinant protein provides an effective system to study the biochemical and physical interactions involved during BoNT binding to nerve cells and protection against its toxicity.     

Complexation of Group IIIA metals with asymmetric tridentate ligands: Synthesis, characterization, formation constants and cytotoxicity

A series of new aluminum(III), gallium(III) and indium(III) complexes with some tridentate Schiff base, viz., N-{pyridine-2-ylmethyl}-2-hydroxy-5-methoxy-benzylideneamine [HL¹], N-{pyridine-2-ylmethyl}-2-hydroxy-benzylideneamine [HL²], N-{pyridine-2-ylmethyl}-2-hydroxy-5-nitro-benzylideneamine [HL³], N-{pyridine-2-ylmethyl}-2-hydroxy-5-bromo-benzylideneamine [HL⁴], N-{pyridine-2-ylethyl}-2-hydroxy-5-methoxy-benzylideneamine [HL⁵], N-{pyridine-2-ylethyl}-2-hydroxy-benzylideneamine [HL⁶], N-{pyridine-2-ylethyl}-2-hydroxy-5-nitro-benzylideneamine [HL⁷], N-{pyridine-2-ylethyl}-2-hydroxy-5-bromo-benzylideneamine [HL⁸], with the general formula [ML₂][Y] (M = Al³⁺, Ga³⁺, In³⁺; Y = NO₃⁻, ClO₄⁻) were synthesised and characterized by elemental analysis, ¹H NMR, FT-IR, UV-Vis spectrophotometry and mass spectrometry. The thermodynamic formation constants of the complexes were determined spectrophotometrically at constant ionic strength (I = 0.10 M NaClO₄) and at 25 °C in methanol. The trend of formation constants of the complexes are as follow:

Al<Ga<In     

Circulating HSPs Levels and Risk of Human Gastrointestinal Related Cancers: A Systematic Review and Meta-analysis

International Journal of Peptide Research and Therapeutics - Heat shock proteins (HSPs) have over-expression in the human malignancies. However, many studies reported inconsistent results. The...     

Antioxidative enzymes activities and accumulation of steroids in hairy roots of Trigonella

Steroidal sapogenins and phytosterols are a group of secondary metabolites which are very considerable in the pharmaceutical industry. Fenugreek (Trigonella foenum-graecum L.) is the good source of these compounds. In recent decades, there is a great interest to production of these compounds by cultivation of transformed roots. In present study, hairy roots induction in two Trigonella species (T. foenum-graeceum, T. monantha) with three strains of Agrobacterium rhizogenes (15,834, A4 and wt) was investigated. Transgenic status of roots was confirmed by PCR using rolB specific primers. Virulence of these strains was examined on explants of leaf, leaf cotyledone and hypocotyle in both species. The best strain was wt for hairy root induction in hypocotyle and leaf explants of T. foenum-graeceum and T. monantha. Significant quantitative differences were showed between shoot, root and hairy roots in both species. Protein content in root and hairy root of both species was significantly lower in comparison with shoot. Superoxide dismutase (SOD) and peroxidase (POX) activities in hairy roots of both species were higher as compared to other organs. The hairy roots of both species showed an ability to synthesize steroidal sapogenins. These results presented that hairy roots could be a suitable procedure for producing sapogenins compounds that have medicinal value in Trigonella.     

Involvement of Hippocampal D1-Like Dopamine Receptors in the Inhibitory Effect of Cannabidiol on Acquisition and Expression of Methamphetamine-Induced Conditioned Place Preference

Cannabidiol (CBD) is a non-psychotomimetic compound with strong potential to decrease the psychostimulant’s rewarding effect with unclear receptors. Furthermore, as a part of the reward circuit, the hippocampus plays a crucial role in regulating the reward properties of drugs as determined by conditioned place preference (CPP). In the current research, CPP was used to evaluate the role of intra-CA1 microinjection of D1-like dopamine receptor antagonists in CBD's inhibitory effect on the acquisition and expression phases of methamphetamine (METH). Animals were treated by METH (1 mg/kg; sc) in a five-day schedule to induce CPP. To find out the impact of D1-like dopamine receptor antagonist, SCH23390, in the CA1 on the inhibitory influence of CBD on the acquisition of METH, the rats received intra-CA1 administration of SCH23390 (0.25, 1, and 4 µg/0.5 µl) following ICV treatment of CBD (10 µg/5 µl) over conditioning phase of METH. Furthermore, animals were given SCH23390 in the CA1 ensuing ICV microinjection of CBD (50 µg/5 µl) in the expression phase of METH to rule out the influence of SCH23390 on the suppressive effect of CBD on the expression of METH CPP. Intra-CA1 microinjection of SCH23390 abolished CBD's suppressive impact on both METH-induced CPP phases without any side effect on the locomotion. The current research disclosed that CBD inhibited the rewarding characteristic of METH via D1-like dopamine receptors in the CA1 region of the hippocampus.

     

Chromoblastomycosis due to Fonsecaea pedrosoi and F. monophora in Cuba

We report two cases of chromoblastomycosis due to Fonsecaea pedrosoi and F. monophora in otherwise healthy Cuban males. Direct microscopic examination of biopsies revealed muriform cells, the hallmark of chromoblastomycosis. The suspected agents were recovered in culture, identified on the basis of morphological criteria and confirmed by sequencing of the internal transcribed spacer regions of rDNA. Final treatment consisted of surgical excision. The patients were successfully cured since there was no relapse after a follow-up of more than a year. In vitro antifungal susceptibility testing of both isolates showed that itraconazole and posaconazole had potent activity. High MICs of amphotericin B (2 μg/ml), fluconazole (>64 μg/ml), anidulafungin (8 μg/ml) and caspofungin (8 μg/ml) were found.