The sympathetic release test: a test used to assess thermoregulation and autonomic control of blood flow

When a subject is heated, the stimulation of temperature-sensitive nerve endings in the skin, and the raising of the central body temperature, results in the reflex release of sympathetic vasoconstrictor tone in the skin of the extremities, causing a measurable temperature increase at the site of release. In the sympathetic release test, the subject is gently heated by placing the feet and calves in a commercially available foot warming pouch or immersing the feet and calves in warm water and wrapping the subject in blankets. Skin blood flow is estimated from measurements of skin temperature in the fingers. Normally skin temperature of the fingers is 65-75 degrees F in cool conditions (environmental temperature: 59-68 degrees F) and rises to 85-95 degrees F during body heating. Deviations in this pattern may mean that there is abnormal sympathetic vasoconstrictor control of skin blood flow. Abnormal skin blood flow can substantially impair an individual's ability to thermoregulate and has important clinical implications. During whole body heating, the skin temperature from three different skin sites is monitored and oral temperature is monitored as an index of core temperature. Students determine the fingertip temperature at which the reflex release of sympathetic activity occurs and its maximal attainment, which reflects the vasodilating capacity of this cutaneous vascular bed. Students should interpret typical sample data for certain clinical conditions (Raynaud's disease, peripheral vascular disease, and postsympathectomy) and explain why there may be altered skin blood flow in these disorders.

Acridine orange leukocyte fluorography in mice

Simultaneous non-invasive visualization of blood vessels and nerves in patients can be obtained in the eye. The retinal vasculature is a target of many retinopathies. Inflammation, readily manifest by leukocyte adhesion to the endothelial lining, is a key pathophysiological mechanism of many retinopathies, making it a valuable and ubiquitous target for disease research. Leukocyte fluorography has been extensively used in the past twenty years; however, fluorescent markers, visualization techniques, and recording methods have differed between studies. The lack of detailed protocol papers regarding leukocyte fluorography, coupled with lack of uniformity between studies, has led to a paucity of standards for leukocyte transit (velocity, adherence, extravasation) in the retina. Here, we give a detailed description of a convenient method using acridine orange (AO) and a commercially available scanning laser ophthalmoscope (SLO, HRA-OCT Spectralis) to view leukocyte behavior in the mouse retina. Normal mice are compared to mice with acute and chronic inflammation. This method can be readily adopted in many research labs.

A curious case of hiccoughs - unusual presentation of eosinophilic granulomatosis with polyangiitis

We report a case of eosinophilic GPA (EGPA) presenting with persistent hiccoughs. The diagnosis was based on clinical and histological findings. Induction treatment with corticosteroids led to clinical improvement and rapid resolution of hiccoughs. Hiccoughing has never before been associated with acute EGPA and may reflect involvement of the phrenic and/or vagus nerves.

Natural Toxicants: Tetrodotoxin

Tetrodotoxin (tetrodotoxin) is a potent neurotoxin, which shuts down electrical signaling in nerves by blocking the voltage-gated sodium channel proteins in nerve cell membranes. It was originally discovered in puffer fish but is found in a range of animal species and thought to be produced by bacteria. The toxin can be lethal to humans being 10 000 times more potent than cyanide. Human fatalities have been attributed to the ingestion of this toxin through consumption of puffer fish, a delicacy in Japan and other regions, and other marine species. The effects of tetrodotoxin poisoning onset quickly and include shortness of breath, numbness, tingling, light-headedness, paralysis, and irregular heartbeat. Treatment usually consists of respiratory assistance as no antidote has been developed. The accepted method of analysis for tetrodotoxin is the mouse bioassay, although recently more ethical assays have been developed including high performance liquid chromatography, biosensor and enzyme-linked immunosorbant assay.

Anatomy and Neurophysiology of Coughing: CHEST Guideline and Expert Panel Report

Bronchopulmonary C-fibers and a subset of mechanically sensitive, acid-sensitive myelinated sensory nerves play essential roles in regulating cough. These vagal sensory nerves terminate primarily in the larynx, trachea, carina and large intrapulmonary bronchi. Other bronchopulmonary sensory nerves, sensory nerves innervating other viscera as well as somatosensory nerves innervating the chest wall, diaphragm and abdominal musculature regulate cough patterning and cough sensitivity. The responsiveness and morphology of the airway vagal sensory nerve subtypes and the extrapulmonary sensory nerves that regulate coughing are described. The brainstem and higher brain control systems that process this sensory information are complex, but our current understanding of them is considerable and increasing. The relevance of these neural systems to clinical phenomena, such as urge to cough and psychological methods for treatment of dystussia, is high and modern imaging methods have revealed potential neural substrates for some features of cough in the human.

Chronic cough as a neuropathic disorder

Chronic cough is a common symptom that can be a daunting challenge for clinicians since treatment of the underlying cause does not always provide adequate relief, an obvious cause can remain elusive, and current antitussives have fairly poor efficacy and undesirable side-effects. Patients with chronic cough typically describe a range of sensory symptoms suggestive of upper-airway and laryngeal neural dysfunction. Additionally, patients often report cough triggered by low-level physical and chemical stimuli, which is suggestive of cough-reflex hyperresponsiveness. Pathophysiological mechanisms underlying peripheral and central augmentation of the afferent cough pathways have been identified, and compelling evidence exists for a neuropathy of vagal sensory nerves after upper-respiratory viral infections or exposure to allergic and non-allergic irritants. In this Personal View, we argue that chronic cough is a neuropathic disorder that arises from neural damage caused by a range of inflammatory, infective, and allergic factors. In support of this idea, we discuss evidence of successful treatment of chronic cough with agents used for treatment of neuropathic pain, such as gabapentin and amitriptyline. Regarding cough as a neuropathic disorder could lead to new, more effective antitussives.

Distribution of terminal nerve entry points to the flexor and extensor groups of forearm muscles:An anatomical study

The motor points of the skeletal muscles, mainly of interest to anatomists and physiologists, have recently attracted much attention from researchers in the field of functional electrical stimulation. The muscle motor point has been defined as the entry point of the motor nerve branch into the epimysium of the muscle belly. Anatomists have pointed out that many muscles in the limbs have multiple motor points. Knowledge of the location of nerve branches and terminal nerve entry points facilitates the exact insertion and the suitable selection of the number of electrodes required for each muscle for functional electrical stimulation. The present work therefore aimed to describe the number, location, and distribution of motor points in the human forearm muscles to obtain optimal hand function in many clinical situations. Twenty three adult human cadaveric forearms were dissected. The numbers of primary nerves and motor points for each muscle were tabulated. The mean numbers and the standard deviation were calculated and grouped in tables. Data analyses were performed with the use of a statistical analysis package (SPSS 13.0). The proximal third of the muscle was the usual part of the muscle that received the motor points. Most of the forearm muscles were innervated from the lateral side and deep surface of the muscle. The information in this study may also be usefully applied in selective denervation procedures to balance muscles in spastic upper limbs. Copyright © 2007 Via Medica.

Morphological study of the uncommon rectus sterni muscle in German cadavers

Objective: The present investigation has been designed to study the incidence of the rectus stern muscle in German human cadavers dissected in the Kingdom of Saudi Arabia, trying to find a postulation for the development of such muscle when present. Design: Gross dissection of 130 cadavers, of both sexes, was performed throughout a period of 10 years. Setting: Department of Anatomy College of Medicine, King Faisal University, Dammam, Saudi Arabia. Intervention: Investigation of the origin and insertion of the rectus sterni and measurements of its length and width. Results: Two adult cadavers, one of each sex, had shown well-developed bilateral rectus stern muscles. All muscles identified were parasternal in position, being superficial to the medial portion of the pectoralis major muscle. Minor morphological differences were observed among the four muscle masses concerning their length, breadth, origin and insertion. Conclusion: The current study has determined the incidence of the rectus sterni muscle, in German cadavers to be 1.54% per bodies examined compared to 4% in cadavers from Saudis. Such a frequency is compared to that reported in different geographic populations. The rectus sterni muscle is innervated by the anterior cutaneous branches of the intercostal nerves. The description of the rectus sterni muscle and its incidence determined in the present study, might be of a great help for clinicians radiographing or tackling the pectoral region.

Bladder interstitial cells:an updated review of current knowledge

The field of bladder research has been energized by the study of novel interstitial cells (IC) over the last decade. Several subgroups of IC are located within the bladder wall and make structural interactions with nerves and smooth muscle, indicating integration with intercellular communication and key physiological functions. Significant progress has been made in the study of bladder ICs' cellular markers, ion channels and receptor expression, electrical and calcium signalling, yet their specific functions in normal bladder filling and emptying remain elusive. There is increasing evidence that the distribution of IC is altered in bladder pathophysiologies suggesting that changes in IC may be linked with the development of bladder dysfunction. This article summarizes the current state of the art of our knowledge of IC in normal bladder and reviews the literature on IC in dysfunctional bladder.

Neural dysfunction following respiratory viral infection as a cause of chronic cough hypersensitivity

Respiratory viral infections are a common cause of acute coughing, an irritating symptom for the patient and an important mechanism of transmission for the virus. Although poorly described, the inflammatory consequences of infection likely induce coughing by chemical (inflammatory mediator) or mechanical (mucous) activation of the cough-evoking sensory nerves that innervate the airway wall. For some individuals, acute cough can evolve into a chronic condition, in which cough and aberrant airway sensations long outlast the initial viral infection. This suggests that some viruses have the capacity to induce persistent plasticity in the neural pathways mediating cough. In this brief review we present the clinical evidence of acute and chronic neural dysfunction following viral respiratory tract infections and explore possible mechanisms by which the nervous system may undergo activation, sensitization and plasticity.

Pulpal Neuropeptide Y levels in relation to pain experience

Neuropeptide Y is a 36 amino acid peptide that belongs to the pancreatic polypeptide family. It co-localises with adrenaline in sympathetic nerves and is released upon sympathetic activation resulting in vasoconstriction. In addition to its vascular effects NPY is also thought to have a role in pain modulation, angiogenesis and immunomodulation. Objectives: The aim of this study was to quantify the levels of NPY in human dental pulp tissue from intact and grossly carious teeth and to relate these results to pain experience. Methods: A total of 48 permanent teeth [mean age 32.1(+/- 11.2 years)] were included in the study, of these 22 were intact and 26 were grossly carious. In the grossly carious group, 17 teeth were reported painful prior to extraction and the remainder were reported non-painful. NPY was measured using a sensitive and specific radioimmunoassay which has been previously described. Pain was scored as either present or absent in all the teeth studied. Results: Of particular interest in this study was the finding that NPY levels were significantly higher in dental pulp tissue from non-painful grossly carious teeth (p= 0.006) compared with painful grossly carious teeth. Conclusions: The increased levels of NPY reported in non-painful grossly carious teeth may suggest a role for NPY in pain modulation in human dental pulp.

Co-expression of NPY and VIP in human dental pulp

Introduction: The regulation of pulpal haemodynamics in health and disease involves sympathetic and parasympathetic mechanisms in which both neuropeptide Y (NPY; a sympathetic vasoconstrictor) and vasoactive intestinal polypeptide (VIP; a parasympathetic vasodilator) may play potential pathophysiological roles. We have previously investigated the levels of NPY or VIP present in human dental pulp tissue and shown that their expression is up-regulated in caries induced pulpal inflammation. Objectives: The aim of this study was to investigate the potential correlation between NPY and VIP levels measured in the same dental pulp samples using radioimmunoassay (RIA). Methods: Pulp tissue was obtained from extracted teeth, classified as follows; healthy (n=22), moderately carious (n=20) and grossly carious (n=26). Samples were processed for RIA by boiling in acetic acid as previously described. The levels of NPY and VIP, measured by RIA, were expressed as ng/gram of pulp tissue. The nature of the relationship between NPY and VIP levels in human pulp tissue was tested by calculating Pearson's product moment correlation coefficient using the linear regression test. Results: Calculation of Pearson product moment correlation coefficient showed a significant negative correlation between NPY and VIP levels in pulp tissue samples from non-carious teeth (p = 0.02, r = -.48). This negative correlation in non-carious teeth changed to a significant positive correlation in carious teeth when the levels of NPY and VIP were compared (p = 0.03, r= 0.311). Conclusions: In non-carious teeth, the negative correlation between NPY and VIP levels is in keeping with the previously described modulatory influence of cholinergic nerves on sympathetic function which may be perturbed as caries develops.

Design, synthesis and validation of an inhibitor of CGRP degradation

Introduction: In addition to their afferent role in detection and signalling noxious stimuli, neuropeptide-containing sensory nerves may initiate and maintain chronic inflammation in diseases such as periodontitis by an efferent process known as neurogenic inflammation. Neuropeptides are susceptible to cleavage by peptidases, and therefore, the exact location and level of expression of peptidases are major determinants of neuropeptide action. Previous studies in our laboratory showed that enzyme components of gingival crevicular fluid (GCF) from periodontitis sites selectively inactivated the neuropeptide calcitonin gene-related peptide (CGRP), known to have a role in inhibiting osteoclastic bone resorption. Objectives: The aim of this study was to design and synthesise a specific inhibitor to prevent the degradation of CGRP by components of GCF. Methods: A hydroxamate-based inhibitor with a biotinylated tag was designed to ensure selectivity for CGRP and ease of use for future purification strategies. The biotinylated peptide hydroxamate contained the P1-P4 amino acid sequence of the potential CGRP cleavage site and was synthesised by solid-phase methods using standard Fmoc chemistry. Inhibition of CGRP metabolism by GCF was determined by MALDI-mass spectrometry (MALDI-MS) using pooled GCF samples from periodontitis patients as a crude source of the CGRP-degrading enzyme. Results: MALDI-MS analysis of CGRP degradation showed almost complete inhibition in the presence of the biotinylated inhibitor. Our results showed that the rate-limiting step in the cleavage of CGRP is endopeptidase cleavage, followed by carboxypeptidase attack. Conclusion: This study demonstrates that the enzyme component of GCF responsible for the degradation of CGRP can be inhibited by a biotinylated hydroxamate modelled on a potential endopeptidase cleavage site. The biotin tag on the inhibitor will facilitate our future purification of the CGRP-cleavage enzyme using a streptavidin-agarose column.

Microneedle Technology

Microneedles (MNs) are minimally invasive devices consisting of numerous micron-sized projections amassed on a baseplate, designed to enhance transdermal drug delivery. When applied to the skin, the needles puncture the outermost layer, the stratum corneum, forming aqueous conduits through which drugs can diffuse to the dermal microcirculation. With an average length of 50-900 μm, MNs are short enough to avoid stimulation of dermal nerves and do not induce bleeding, yet gain access to the skin's rich microcirculation for drug delivery. MNs have been extensively investigated for drug and vaccine delivery, demonstrating their efficacy at increasing the number of compounds amenable to delivery through the skin. This chapter discusses the materials and fabrication methods involved in MN production, alongside the different types of MN arrays and their delivery capabilities. The field has expanded to consider novel applications of MNs including minimally invasive patient monitoring, ocular delivery and enhanced administration of cosmeceuticals. Patient usage and effects on the skin are also considered in terms of safety, efficacy and acceptability. The next steps in MN development are to focus on the scale-up of manufacturing processes, a challenge considering the number of small-scale methods detailed in the literature. Regulatory guidance is awaited to direct this, alongside provision of clearer patient instruction for safe and effective use of MN devices. MNs have tremendous potential to yield real benefits for patients and industry and with continued research in the key areas highlighted, this will begin to be realised over the next number of years.

Salivary proteomics and peptidomics of type 1 diabetes Mellitus

A Diabetes Mellitus (DM) compreende um conjunto de desordens metabólicas comuns caracterizadas por hiperglicemia, que afeta diferentes órgãos do organismo. Ao longo do tempo, ocorrem danos microvasculares no glomérulo renal, retina e nervos periféricos, bem como doença macrovascular nas artérias. A composição da saliva também é afetada pela DM, com consequências na homeostasia oral. No entanto, o proteoma e o peptidoma salivar têm sido pouco explorados na DM tipo 1 e nas suas complicações crónicas. Tendo em conta o crescente interesse na saliva como fluido diagnóstico, o objetivo principal deste trabalho foi avaliar os eventos proteolíticos subjacentes à DM tipo 1 e às suas complicações microvasculares, bem como, caracterizar as alterações induzidas pela DM tipo 1 no proteoma e peptidoma salivar. A DM tipo 1 e particularmente as complicações microvasculares associadas modulam o perfil proteolítico dos fluidos biológicos, com diferenças significativas de atividade observadas na urina e saliva, atribuídas principalmente ao complexo Metaloproteinase da Matriz (MMP)-9/lipocalina associada à gelatinase de neutrófilos, aminopeptidase N, azurocidina e calicreína 1. O aumento da atividade proteolítica observado na saliva total dos diabéticos resultou no aumento da percentagem de péptidos, principalmente de um número acrescido de fragmentos de colagénio do tipo I, refletindo possivelmente um estado inflamatório crónico dos tecidos orais e periodontais. O peptidoma também corrobora uma maior suscetibilidade das proteínas salivares, especificamente, das proteínas ricas em prolina básicas (bPRP) 1, bPRP2 e proteínas ricas em prolina ácidas (aPRP) à proteólise, evidenciando a geração de fragmentos de proteínas associadas à ligação a bactérias. A análise do proteoma salivar baseada em iTRAQ mostrou uma sobre-expressão de L-plastina, fator do adenocarcinoma do pâncreas e das proteínas S100-A8 e S100-A9, enfatizando a importância do sistema imune inato na patogénese da DM tipo 1 e das complicações microvasculares associadas. A análise integrada de todas as proteínas expressas diferencialmente entre os pacientes diabéticos com ou sem complicações microvasculares e indivíduos saudáveis foi realizada com o STRING, onde se observam três conjuntos funcionalmente ligados, um compreende a interação entre o colagénio tipo I, colagénio tipo II e MMP-9, um segundo conjunto envolve a MMP-2 e o colagénio de tipo I e um terceiro conjunto composto por proteínas salivares e inflamatórias. Estes conjuntos estão associados com as vias Kegg de interação recetor-matriz extracelular, de adesão focal e migração transendotelial dos leucócitos. Por outro lado, a análise do proteoma e peptidoma salivar destacou potenciais biomarcadores para o diagnóstico e prognóstico da DM tipo 1 e das suas complicações.