Sensitive Afferents Research Articles

Selective Inhibitory Effect of Imidafenacin and 5-Hydroxymethyl Tolterodine on Capsaicin Sensitive C Fibers of the Primary Bladder Mechanosensitive Afferent Nerves in the Rat

Imidafenacin and fesoterodine are used to treat overactive bladder. Imidafenacin, fesoterodine and its active metabolite 5-hydroxymethyl tolterodine are muscarinic receptor antagonists. It is believed that these agents act on afferent nerves in addition to smooth muscle. We investigated the effects of imidafenacin and 5-hydroxymethyl tolterodine on single unit afferent activity of mechanosensitive capsaicin sensitive and insensitive primary bladder afferent nerve fibers in rats. Female Sprague Dawley® rats were anesthetized. Single unit afferent activity was recorded from the L6 dorsal roots and classified by conduction velocity as that of Aδ or C fibers. After measuring control single afferent activity during constant filling cystometry the procedure was repeated with intravenous administration of imidafenacin (0.3 to 30 μg/kg) or 5-hydroxymethyl tolterodine (0.01 to 1 mg/kg) at cumulative doses with or without intravesical capsaicin or oxotremorine-M instillation. A total of 139 single unit afferent fibers were isolated from 111 rats, including 19 Aδ and 120 C fibers. Neither imidafenacin nor 5-hydroxymethyl tolterodine significantly affected the overall single unit afferent activity of Aδ or C fibers. Based on capsaicin sensitivity C fibers were divided into capsaicin sensitive and insensitive groups. Each antimuscarinic inhibited the single unit afferent activity of capsaicin sensitive C fibers but not of capsaicin insensitive C fibers at the highest dose. Moreover, oxotremorine-M facilitated single unit afferent activity in a proportion of C fibers. The facilitated single unit afferent activity was significantly attenuated by the highest dose of imidafenacin. These findings demonstrate that imidafenacin and 5-hydroxymethyl tolterodine can selectively inhibit capsaicin sensitive C fibers among mechanosensitive bladder afferents by antagonizing bladder muscarinic receptors.

Neurophysiology of the esophagus.

The following, from the 12th OESO World Conference: Cancers of the Esophagus, includes commentaries on the methods and characteristics of esophageal afferents in humans; the pitfalls in characterization of mechanosensitive afferents; the sensitization of esophageal afferents in human studies; the brain source modeling in the understanding of the esophagus-brain axis; the use of evoked brain potentials in the esophagus; and measuring descending inhibition in animal and human studies.

Targeting TRP Channels For Novel Migraine Therapeutics

Migraine is increasingly understood to be a disorder of the brain. In susceptible individuals, a variety of “triggers” may influence altered central excitability, resulting in the activation and sensitization of trigeminal nociceptive afferents surrounding blood vessels (i.e., the trigeminovascular system), leading to migraine pain. Transient receptor potential (TRP) channels are expressed in a subset of dural afferents, including those containing calcitonin gene related peptide (CGRP). Activation of TRP channels promotes excitation of nociceptive afferent fibers and potentially lead to pain. In addition to pain, allodynia to mechanical and cold stimuli can result from sensitization of both peripheral afferents and of central pain pathways. TRP channels respond to a variety of endogenous conditions including chemical mediators and low pH. These channels can be activated by exogenous stimuli including a wide range of chemical and environmental irritants, some of which have been demonstrated to trigger migraine in humans. Activation of TRP channels can elicit CGRP release, and blocking the effects of CGRP through receptor antagonism or antibody strategies has been demonstrated to be effective in the treatment of migraine. Identification of approaches that can prevent activation of TRP channels provides an additional novel strategy for discovery of migraine therapeutics.

Deletion of interleukin-6 signal transducer gp130 in small sensory neurons attenuates mechanonociception and down-regulates TRPA1 expression.

Glycoprotein 130 (gp130) is the signal transducing receptor subunit for cytokines of the interleukin-6 (IL-6) family, and it is expressed in a multitude of cell types of the immune and nervous system. IL-6-like cytokines are not only key regulators of innate immunity and inflammation but are also essential factors for the differentiation and development of the somatosensory system. Mice with a null mutation of gp130 in primary nociceptive afferents (SNS-gp130(-/-)) are largely protected from hypersensitivity to mechanical stimuli in mouse models of pathological pain. Therefore, we set out to investigate how neuronal gp130 regulates mechanonociception. SNS-gp130(-/-) mice revealed reduced mechanosensitivity to high mechanical forces in the von Frey assay in vivo, and this was associated with a reduced sensitivity of nociceptive primary afferents in vitro. Together with these findings, transient receptor potential ankyrin 1 (TRPA1) mRNA expression was significantly reduced in DRG from SNS-gp130(-/-) mice. This was also reflected by a reduced number of neurons responding with calcium transients to TRPA1 agonists in primary DRG cultures. Downregulation of Trpa1 expression was predominantly discovered in nonpeptidergic neurons, with the deficit becoming evident during stages of early postnatal development. Regulation of Trpa1 mRNA expression levels downstream of gp130 involved the classical Janus kinase family-signal transducer and activator of transcription pathway. Our results closely link proinflammatory cytokines to the expression of TRPA1, both of which have been shown to contribute to hypersensitive pain states. We suggest that gp130 has an essential role in mechanonociception and in the regulation of TRPA1 expression.

Vagal leptin signalling: A double agent in energy homeostasis?

In this issue of Molecular Metabolism, de Lartigue et al. reveal the effect of leptin receptor depletion in vagal afferents on food intake and weight regulation. de Lartigue et al. showed deletion of leptin receptor in Nav1.8 expressing sensory neurons resulted in increased food intake (specifically in the dark phase), reduced sensitivity to the satiety hormone cholecystokinin (CCK), which ultimately led to increased adiposity and a modest increase in weight. Curiously, feeding this knockout line a high fat diet led to no further weight gain, compared to the chow fed knockout animals, and, in fact, they were substantially lighter than high fat diet fed wildtype mice. This landmark paper provides direct evidence for the physiological importance of leptin–vagal afferent interactions in satiety signalling and long term weight regulation. Currently, the long term regulation of energy homeostasis is largely attributed to homeostatic pathways within the central nervous system. The satiety hormone, leptin, secreted primarily from white adipose tissue, provides a signal to the hypothalamus reflecting the size of fat stores [1,2] and serves as a long-term regulator of nutrient intake, adiposity and body weight [3]. However, although the levels of circulating leptin reflect the degree of adiposity [1], in obesity, where leptin levels are high, there is an inability for leptin to suitably regulate body weight, a phenomenon dubbed ‘leptin resistance’. In the periphery, it is well recognised that signals generated in the gastrointestinal tract are involved in the short term regulation of food intake influencing meal size and duration [4]. This involves integration of both gastric and small intestine feedback. In the small intestine this signalling is thought to be an enteroendocrine response to nutrients, and the data of de Lartigue et al. indicate that leptin receptor disruption, specifically in vagal afferents, decreases vagal afferent sensitivity to the anorexigenic hormone CCK, thus presumably reducing the satiety signal arising from the small intestine. On the other hand, CCK also activates gastric afferents [5] and, as there were no means to actually determine the location of the afferent ending involved this may be somewhat premature. Additionally, the authors fail to consider satiety signals from the stomach. In the stomach, vagal afferent satiety signals are generated largely in response to mechanical stimuli. Gastric vagal afferents can also be acted on by a number of appetite regulating peptides, including leptin, to regulate food intake [6], suggesting the results of de Lartigue et al. likely have intestinal and gastric contributions. Although gastrointestinal vagal afferents are an important link between the periphery and the central nervous system the influence of vagal afferent satiety signals on long term regulation of energy homeostasis has been under debate for many years. This is largely due to the multiple central and peripheral sites of action of appetite regulating hormones (Figure 1). By specifically manipulating leptin signalling in vagal afferents de Lartigue et al. have established conclusively that vagal afferents are an important component to regulate satiety and influence long term food intake and weight gain. Figure 1 Illustration of the effect of leptin on appetite regulation within the central nervous system and on peripheral vagal afferents under standard and high fat diet induced obese conditions. An interesting observation in the data of de Lartigue et al. is that in high fat diet conditions weight gain is reduced in vagal afferent leptin receptor knockout mice. This implies that in high fat diet conditions leptin has ‘orexigenic’ effects possibly increasing food intake. Unfortunately, the authors fail to provide food intake data on this group of mice and therefore at this stage this cannot be confirmed. However, a previous report [7] indicating a ‘switch’ in the effect of leptin on gastric vagal afferents in high fat diet conditions supports the data of de Lartigue et al. Thus this data introduces the intriguing possibility that leptin is a ‘double agent’ normally having anorexigenic effects but under certain physiological/nutritional states switching to an orexigenic agent. As a whole de Lartigue et al. demonstrate the importance of vagal afferents in regulation of long term food intake and weight with vagal leptin signalling required to maintain a healthy weight, but in contrast, are also required to develop high fat diet induced obesity. It is apparent that a better understanding of the underlying mechanisms in vagal afferent satiety signalling under varying states of nutrition is essential before any pharmacotherapy for the treatment of obesity can be established.

Multiple roles for NaV1.9 in the activation of visceral afferents by noxious inflammatory, mechanical, and human disease–derived stimuli

Chronic visceral pain affects millions of individuals worldwide and remains poorly understood, with current therapeutic options constrained by gastrointestinal adverse effects. Visceral pain is strongly associated with inflammation and distension of the gut. Here we report that the voltage-gated sodium channel subtype NaV1.9 is expressed in half of gut-projecting rodent dorsal root ganglia sensory neurons. We show that NaV1.9 is required for normal mechanosensation, for direct excitation and for sensitization of mouse colonic afferents by mediators from inflammatory bowel disease tissues, and by noxious inflammatory mediators individually. Excitatory responses to ATP or PGE2 were substantially reduced in NaV1.9−/− mice. Deletion of NaV1.9 substantially attenuates excitation and subsequent mechanical hypersensitivity after application of inflammatory soup (IS) (bradykinin, ATP, histamine, PGE2, and 5HT) to visceral nociceptors located in the serosa and mesentery. Responses to mechanical stimulation of mesenteric afferents were also reduced by loss of NaV1.9, and there was a rightward shift in stimulus–response function to ramp colonic distension. By contrast, responses to rapid, high-intensity phasic distension of the colon are initially unaffected; however, run-down of responses to repeat phasic distension were exacerbated in NaV1.9−/− afferents. Finally colonic afferent activation by supernatants derived from inflamed human tissue was greatly reduced in NaV1.9−/− mice. These results demonstrate that NaV1.9 is required for persistence of responses to intense mechanical stimulation, contributes to inflammatory mechanical hypersensitivity, and is essential for activation by noxious inflammatory mediators, including those from diseased human bowel. These observations indicate that NaV1.9 represents a high-value target for development of visceral analgesics.

Abstracts fromThe 32nd AnnualNational Neurotrauma SymposiumJune 29–July 2, 2014San Francisco, California

Abstracts fromThe 32^nd AnnualNational Neurotrauma SymposiumJune 29–July 2, 2014San Francisco, California

Vertebrate galectins: endogenous regulators of ionotropic glutamate receptors?

Lectins are proteins that bind specific carbohydrate residues on glycoproteins and glycolipids. In many cases, such binding alters the functional properties of the targeted protein or lipid and triggers a change in cell physiology. First characterized in plants, lectins with diverse carbohydrate specificity have subsequently been identified in animals, where they serve a variety of different functions including both intracellular regulation of protein sorting and extracellular interactions with carbohydrate moieties on the cell surface and in the extracellular matrix (Dodd & Drickamer, 2001). Both plant and animal lectins typically exist as dimers or multimers with their biological action being dependent on, or enhanced by, the ability to form cross links between protein subunits. In the case of ionotropic glutamate receptors (iGluRs) early work with plant lectins demonstrated a potentiating action on agonist-evoked currents recorded in insect and vertebrate cells. Vertebrates express three distinct iGluR families, but only α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and kainate (KA) subtypes exhibit strong direct modulation by lectins while N-methyl-d-aspartate (NMDA) receptors are relatively unaffected. Brief exposure to wheat germ agglutinin, concanavalin A (ConA) or isolectin B4, with specificity for N-acetylglucosamine, mannose and β-galactosides, respectively, increases the amplitude of steady-state currents evoked via KA and/or AMPA receptors (Mayer & Vyklicky, 1989; Huettner, 1990). Now, a study from Geoff Swanson's lab in this issue of The Journal of Physiology (Copits et al. 2014) investigates the ability of vertebrate galectins, including recombinant human galectin-1, and congerin I and II from the conger eel, to regulate native and recombinant AMPA and KA receptors. Galectins bind specifically to β-galactoside glycoconjugates and the three galectins studied by Copits et al. (2014) can each form a bivalent homodimer. In contrast to tetravalent ConA, which substantially increases currents mediated by most AMPA and KA receptor isoforms, galectin application produced a remarkable diversity of effects on recombinant iGluRs depending on the specific subunits that were expressed and on which galectin was applied. For example, congerin caused a substantial slowing of AMPA receptor desensitization with an increase in steady-state current in cells expressing homomeric GluA4 and some of the cells expressing GluA1 whereas human galectin-1 had a greater effect on GluA1 than GluA4. In addition, congerin, but not galectin-1, reduced peak current with little change in kinetics in cells that expressed homomeric GluA2 or the GluA1/GluA2 heteromeric combination. Similarly, both eel and human galectins elicited a significantly greater slowing of desensitization in homomeric GluK2 KA receptors than homomeric GluK1 or heteromeric GluK2/GluK5 combinations. Modulation persisted in heterologous cells that were co-transfected with AMPA or KA receptor subunits together with their auxiliary subunits stargazin or Neto2, respectively. Mechanistically, galectin modulation of GluK2 was reduced by drugs that interfere with oligosaccharide processing and prevented by point mutations that eliminate three essential glycosylation sites at the interface between the receptor's amino terminal and ligand binding domains (Everts et al. 1999). In addition, GluK2 modulation was enhanced by linkers that stabilized the recombinant galectin as an obligate dimer, which is consistent with the idea that iGluR modulation involves crosslinking subunits via their attached glycoconjugates. In future work, it would be interesting to determine whether galectin–3, which can assemble into a pentameric oligomer, might elicit even stronger modulation. When tested on neuronal AMPA receptors in cultures prepared from rat hippocampus, congerin failed to change the amplitude or time course of agonist-evoked or spontaneous excitatory synaptic currents, possibly owing to additional modifications of glycoconjugates in central neurons that render them resistant to galectins or, alternatively, prior occupation of the binding site by endogenous lectins produced in the cultures that occlude the binding of acutely applied exogenous galectin. On the other hand, both congerin and human galectin-1 slowed desensitization of KA receptor-mediated currents recorded in freshly dissociated mouse sensory neurons, cells that are known to express galectin-1 and -3 as well as β-galactoside glycoconjugates (Jessell et al. 1990). Thus, Swanson and colleagues (Copits et al. 2014) have established the potential for native receptors to be modulated by endogenous lectins, but much work remains to determine where and when such regulation actually occurs. Mice lacking galectin-1 are viable but exhibit changes in primary afferent morphology and sensitivity (McGraw et al. 2005) as well as resistance to death of central neurons subsequent to pilocarpine-induced seizures (Bischoff et al. 2012). Whether or not these phenotypes involve alteration in iGluR modulation remains to be determined, but the availability of both galectin–1 and galectin–3 knock-outs should aid in unravelling their potential role in regulating excitatory amino acid receptors.

The increased distensibility of the wall of cerebral arterial network may play a role in the pathogenic mechanism of migraine headache

The aim was to evaluate whether patients with episodic migraine with (MA+) and without aura (MA-), during the interictal period of migraine would have an altered distensibility of the wall of cerebral arterial network and whether it would play a role in migraine headache. To evaluate the distensibility of the wall of cerebral arterial network, we measured the time-delay in milliseconds (ms) between the R-wave of an electrocardiogram and the arterial pulse wave of cerebral microcirculation (R-APWCMtd) on the frontal cortex detected by near-infrared spectroscopy (NIRS) in 10 patients with MA+ (age 39.5 ± 12.2 years), in 10 with MA- (age 40.3 ± 10.2 years), according to ICHD-3 criteria 2012, during the interictal period of migraine, and in 15 age-, sex- and height-matched healthy control subjects. The patients with migraine had a significantly longer R-APWCMtd than the control subjects F = 13.4, p < 0.001: MA+:+38.3 ms; MA-:+34.7 ms indicating an increased distensibility of the wall of cerebral arterial network. In multiple regression analysis, R-APWCMtd was significantly associated with migraine (R (2) = 0.50, p < 0.0001) but not with age, gender, height, migraine attack frequency and disease duration. The increased distensibility leads to an increased flow pulsatility into intracranial dural meningeal vessels that may lead to a mechanical stimulation of the nociceptors that innervate the dural vasculature. This condition may play a role in promoting the sensitization of trigeminovascular afferents and sterile inflammation within the dura mater that are fundamental to the pathogenesis of migraine headache.

Mo2036 Remodeling of Colonic Sensory Afferents in a Rat Model of Post-Infectious Gut Dysfunction: Implication of Neurotrophic Factors

Background: Inflammation-induced visceral hypersensitivity is associated, at least partially, to peripheral sensitization of sensory afferents by locally released neurotrophic factors. Aims: Here, we explore the potential role of neurotrophic factors in long-term changes in visceral afferents innervating non-primary areas of inflammation (colon) in a rat model of Trichinella spiralis (TS)-induced gut dysfunction. Methods: Inflammatory-like changes, mucosal mast cells (MMCs) dynamics, and expression of nerve growth factor (NGF) and glial cell linederived neurotrophic factors (GDNF, artemin and neurturin) were determined in the colon of control and TS-infected rats, up to day 30 post-infection (PI). Markers of sensory afferents (high affinity nerve growth factor receptor, TrkA; GDNF family receptor alpha 3, GFRα3; calcitonin gene-related peptide, CGRP; and transient receptor potential vanilloid channel1, TRPV1) were assessed in thoracolumbar (T12-L2, TL) and lumbosacral (L6-S2, LS) DRGs. Functionality of colonic afferents at 14 and 30 days PI was determined by assessing changes in TRPV1 levels in TL and LS DRGs following intracolonic capsaicin. Results: TS infection induced a primary jejunitis with concomitant inflammatory-like changes in the colon: an increase in histological scores, IL-13 up-regulation and MMC infiltration (Table 1). Inflammation was partly resolved at day 30 PI, except for a persistent MMC infiltrate (Table 1). Colonic expression of neurotrophic factors showed a transient reduction by day 14 PI (25%50%), with the exception of artemin that showed a biphasic response with an up-regulation (by 30%; P<0.05) at day 30 PI (Table 2). Overall, TS infection did not affect DRG expression of none of the genes assessed, except for a transient down-regulation of TPRV1 in TL DRGs at day 14 PI. Afferent stimulation with intracolonic capsaicin induced a down-regulation of TRPV1 expression in TL and LS DRGs, an effect significantly enhanced in LS DRGs of TSinfected rats, at either 14 or 30 days PI (Table 2). Intracolonic capsaicin also up-regulated peripheral artemin in non-infected rats, an effect absent in TS-infected animals at day 30 PI (Table 2). Capsaicin-induced changes in colonic expression of artemin correlated negatively with the down-regulation of TRPV1 in LS DRGs. Conclusions: During intestinal inflammation, changes in expression of neurotrophic factors and remodeling of visceral afferents are observed outside the primary region affected by the inflammatory insult. These observations suggest that extended remodelation of sensory afferents, probably related to local changes in neurotropins, might occur during inflammatory conditions of the gut. Similar mechanisms might be operating in states of widespread alterations of visceral sensitivity. Table 1 Colonic inflammation-related changes during T. spiralis infection

The sensory innervation of the calvarial periosteum is nociceptive and contributes to headache-like behavior

Headaches are thought to result from the activation and sensitization of nociceptors that innervate deep cephalic tissues. A large body of evidence supports the view that some types of headaches originate intracranially, from activation of sensory neurons that innervate the cranial meninges. However, the notion of an extracranial origin of headaches continues to be entertained, although the identity of deep extracranial cephalic tissues that might contribute to headaches remains elusive. Here we employed anatomical, electrophysiological, and behavioral approaches in rats to test the hypothesis that the sensory innervation of the calvarial periosteum is nociceptive. Neural tracing indicated that the calvarial periosteum overlying the frontal and parietal bones is innervated primarily by small and medium-sized neurons in the trigeminal ganglion’s ophthalmic division. In vivo single-unit recording in the trigeminal ganglion revealed that calvarial periosteal afferents have slowly conducting axons, are mechanosensitive, and respond to inflammatory mediators, consistent with a nociceptive function. Two distinct neuronal populations were distinguished based on their peripheral axonal trajectory: one that reached the periosteum through extracranial branches of the trigeminal nerve, and another that took an intracranial trajectory, innervating the cranial dura and apparently reaching the periosteum via the calvarial sutures. In behavioral studies, inflammatory stimulation of these afferents promoted periorbital tactile hypersensitivity, a sensory change linked to primary headaches. Activation and sensitization of calvarial periosteal afferents could play a role in mediating primary headaches of extracranial and perhaps also intracranial origin, as well as secondary headaches such as postcraniotomy and posttraumatic headaches. Targeting calvarial periosteal afferents may be effective in ameliorating these headaches.

Neural basis of new clinical vestibular tests: otolithic neural responses to sound and vibration.

Extracellular single neuron recording and labelling studies of primary vestibular afferents in Scarpa's ganglion have shown that guinea-pig otolithic afferents with irregular resting discharge are preferentially activated by 500Hz bone-conducted vibration (BCV) and many also by 500Hz air-conducted sound (ACS) at low threshold and high sensitivity. Very few afferent neurons from any semicircular canal are activated by these stimuli and then only at high intensity. Tracing the origin of the activated neurons shows that these sensitive otolithic afferents originate mainly from a specialized region, the striola, of both the utricular and saccular maculae. This same 500Hz BCV elicits vestibular-dependent eye movements in alert guinea-pigs and in healthy humans. These stimuli evoke myogenic potentials, vestibular-evoked myogenic potentials (VEMPs), which are used to test the function of the utricular and saccular maculae in human patients. Although utricular and saccular afferents can both be activated by BCV and ACS, the differential projection of utricular and saccular afferents to different muscle groups allows for differentiation of the function of these two sensory regions. The basic neural data support the conclusion that in human patients in response to brief 500Hz BCV delivered to Fz (the midline of the forehead at the hairline), the cervical VEMP indicates predominantly saccular function and the ocular VEMP indicates predominantly utricular function. The neural, anatomical and behavioural evidence underpins clinical tests of otolith function in humans using sound and vibration.

TRPA1 mediates bladder hyperalgesia in a mouse model of cystitis

Urinary bladder pain is a primary symptom associated with interstitial cystitis/painful bladder syndrome. We used systemic injections of cyclophosphamide (CYP), an alkylating antineoplastic agent, to induce cystitis and examine the roles of 2 channels previously demonstrated to be required for inflammatory visceral hyperalgesia: transient receptor potential vanilloid-1 (TRPV1) and ankyrin-1 (TRPA1). Injection of CYP (100mg/kg, i.p.) every other day for 5days was accompanied by bladder edema and urothelial ulceration, but without significant plasma extravasation or infiltration of neutrophils. Toluidine blue staining showed a significant increase in the number of degranulated bladder mast cells after CYP treatment. Despite this mild pathology, CYP-treated mice exhibited bladder hyperalgesia 1 day after the final injection that persisted 7days later. Although many previous studies of visceral hyperalgesia have reported changes in dorsal root ganglion neuron TRPV1 expression and/or function, we found no change in bladder afferent TRPV1 expression or sensitivity on the basis of the percentage of bladder afferents responsive to capsaicin, including at submaximal concentrations. In contrast, the percentage of bladder afferents expressing functional TRPA1 protein (ie, those responsive to mustard oil) increased ∼2.5-fold 1day after CYP treatment, and remained significantly elevated 7days later. Moreover, bladder hyperalgesia was reversed by acute treatment with the TRPA1 antagonist HC-030031 (300mg/kg, i.p.). Our results indicate that CYP-induced bladder hyperalgesia can be induced without robust inflammation or changes in primary afferent TRPV1. However, significant changes were observed in TRPA1 expression, and blockade of TRPA1 alleviated CYP-induced bladder hyperalgesia.

Interactive effects of trigeminal nerve stimulation and muscle metaboreflex activation on muscle sympathetic nerve activity in healthy humans (1170.5)

We investigated the independent and interactive effects of trigeminal cutaneous receptor stimulation and activation of metabolically sensitive skeletal muscle afferents (muscle metaboreflex) on muscle sympathetic nerve activity (MSNA). Heart rate (HR; ECG), blood pressure (BP; Finometer) and MSNA (microneurography) were continuously recorded in eight healthy men (29±2 yr) during three experimental conditions; 1) facial cooling [0°C, trigeminal stimulation, TGS], 2) muscle metaboreflex activation by post‐exercise ischemia [PEI] following isometric handgrip at 30% maximum voluntary contraction, and 3) simultaneous TGS and PEI. Trials were counterbalanced and respiratory rate and tidal volume controlled at baseline, TGS, PEI and TGS+PEI. TGS produced a significant decrease in HR (Δ‐5±2 bpm; mean±SE), and increase in mean BP (Δ14±3 mmHg) and MSNA total activity (Δ353±118% AU). During PEI, HR remained at resting levels (Δ3±2 bpm; mean±SE), while mean BP (Δ25±3 mmHg) and MSNA total activity (Δ234±83% AU) increased. Finally, during combined TGS and PEI, HR was unchanged (Δ0±2 bpm; mean±SE), while mean BP (Δ30±3 mmHg) and MSNA total activity (Δ331±100% AU) were increased. Thus, excitatory inputs from the trigeminal nerve and metabolically sensitive skeletal muscle afferents do not appear to summate algebraically in determining a MSNA response, but rather exhibit neural occlusion.Grant Funding Source: Supported by CAPES, CNPq, FAPERJ.

Group III/IV muscle afferents are important for the pressor response to dynamic exercise in healthy humans (1165.9)

The onset of dynamic exercise typically exhibits a triphasic blood pressure (BP) response, consisting of an initial rise (0‐3s) followed by a transient drop below resting levels (4‐10s) and then a sustained increase (&gt;10s). We investigated the role of skeletal muscle afferents in modulating the dynamics of BP at the onset of exercise. Beat‐to‐beat BP (brachial artery), heart rate (HR), stroke volume (SV), cardiac output (CO) were continuously monitored in 8 healthy men (28±3 yr) while performing leg cycling exercise (80W) under control (CTRL) and lumbar intrathecal fentanyl (FENT) (impairing cephalad projection of opioid receptor sensitive muscle afferents). To evaluate whether cephalad migration of fentanyl affected cardiovascular control centers, we assessed the pressor response to handgrip exercise in both conditions. At the exercise onset, mean BP increased from baseline in the first 3s (+5±1 mmHg, P&lt;0.05) and decreased to a nadir at 10s (‐4±3 mmHg, P&lt;0.05). Fentanyl did not affect BP response during the first 10s, but blunted its increase thereafter (CTRL: +9±3 vs. FENT: 0±2 mmHg at 50s; P&lt;0.05). HR, SV and CO responses to exercise were not affected by fentanyl. Similar mean BP responses to static handgrip in both conditions indicated that cephalad migration of fentanyl was avoided. Our findings demonstrate an important contribution of group III/IV muscle afferent fibers to BP response to leg cycling exercise.Grant Funding Source: Supported by the Brazilian Science Without Borders Program

Spinal μ‐opioid receptor sensitive locomotor muscle afferents decrease responsiveness of motor cortical cells and diminish cortical voluntary activation (705.1)

We investigated the influence of group III/IV locomotor muscle afferents on the development of supraspinal fatigue and changes in motor cortical cell responsiveness by comparison of cortical versus spinal excitability during sustained cycling exercise. Six males performed constant‐load cycling exercise (80% peak power‐output) to exhaustion under control conditions (CTRL) and with lumbar intrathecal fentanyl (FENT) impairing feedback from µ‐opioid receptor‐sensitive lower limb muscle afferents. Pre‐to‐post exercise changes in voluntary activation (VA) and potentiated quadriceps twitch force (∆Qtw) were assessed via transcranial magnetic (TMS) and electrical femoral nerve stimulation (FNS). During cycling, vastus lateralis responses evoked by TMS (MEP) and electrical cervicomedullary stimulation (CMEP) were normalized to FNS‐evoked M‐waves. Although maximum voluntary quadriceps torque was similarly reduced following both trials (~14%; P&lt;0.05), ∆Qtw was greater in FENT vs. CTRL (51 ± 5% vs. 62 ± 7%; P&lt;0.05). Furthermore, VA was reduced by 6% in CTRL (P&lt;0.05) but remained unchanged in FENT. MEP increased throughout the exercise in FENT (14 ± 5%; P&lt;0.05) but remained unchanged in CTRL. CMEP did not change in either session. Our findings suggest that group III/IV muscle afferents compromise motor cortical cell responsiveness during locomotor exercise and facilitate supraspinal fatigue.Grant Funding Source: HL_103786; HL‐116579

MP76-06 URINARY NEUROTROPHINS AND QMAX VARIATION MAY PREDICT DE NOVO URGENCY IN SUI PATIENTS AFTER A MIDURETHRAL SLING (MUS) SURGERY

You have accessJournal of UrologyUrodynamics/Incontinence/Female Urology: Non-neurogenic Voiding Dysfunction1 Apr 2014MP76-06 URINARY NEUROTROPHINS AND QMAX VARIATION MAY PREDICT DE NOVO URGENCY IN SUI PATIENTS AFTER A MIDURETHRAL SLING (MUS) SURGERY Tiago Antunes-Lopes, Daniel Costa, Rui Pinto, Sérgio Carvalho-Barros, Carlos Martins-Silva, Célia Duarte-Cruz, and Francisco Cruz Tiago Antunes-LopesTiago Antunes-Lopes More articles by this author , Daniel CostaDaniel Costa More articles by this author , Rui PintoRui Pinto More articles by this author , Sérgio Carvalho-BarrosSérgio Carvalho-Barros More articles by this author , Carlos Martins-SilvaCarlos Martins-Silva More articles by this author , Célia Duarte-CruzCélia Duarte-Cruz More articles by this author , and Francisco CruzFrancisco Cruz More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2014.02.2399AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Nearly 10% of women with stress urinary incontinence (SUI) submitted to a midurethral sling (MUS) surgery develop urgency, eventually maintaining incontinence and low quality of life. High levels of nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) were measured in the urine of OAB patients, while low levels of both neurotrophins (NT) were reported in SUI women. Therefore, NT might be involved in the emergence of urgency, throughout sensitization of bladder afferent nerves. We measured urinary levels of NGF and BDNF, and calculated maximum flow rate (Qmax) variation, in women with SUI, before and after a midurethral sling (MUS) surgery, to investigate the role of NT and flow reduction in the emergence of urgency. METHODS Urine samples were collected from 31 patients with pure clinical SUI and 20 healthy women. SUI patients underwent surgical treatment with a transobturator MUS or a mini-sling. At 1-yr follow-up, patients were reevaluated and the onset of de novo urgency was investigated. Uroflowmetry and urine sampling were repeated. Urinary NGF and BDNF levels were measured by ELISA and normalized to creatinine concentration (pg/mg). The Qmax variation was calculated for each patient and correlated with urinary levels of NT after surgery. RESULTS At baseline, urinary levels of NGF and BDNF were similar between SUI and healthy women (NGF/Cr: 199.0±176.5 vs. 188.3±290.2, p=0.884; BDNF/Cr: 173.8±193.4 vs. 110.4±159.5, p=0.236). In SUI patients, 1-yr after surgery, NGF increased to 448.2±360.3 (vs. baseline, p=0.002) and BDNF to 472.8±541.8 (vs. baseline, p=0.002). Eight patients developed de novo urgency. In this group, NGF increased from 183.5±182.3 to 646.7±471.9 (p=0.034), and BDNF from 188.3±159.0 to 815.6±833.7 (p=0.037). In patients without urgency (n=23), there was a lower, but significant, increase of NT (NGF: 204.3±178.2 to 379.2±294.8, p=0.031; BDNF: 177.0±206.5 to 352.5±348.6, p=0.021). Comparing both groups, patients who developed urgency had significantly higher levels of NGF (646.7±471.9 vs. 379.2±294.8, p=0.029) and BDNF (815.6±833.7 vs. 352.5±348.6, p=0.034). Moreover, women who developed urgency had a higher Qmax reduction than women without urgency (23.8±4.9% vs. 18.3±8.4%, p=0.085). CONCLUSIONS In SUI women, after a MUS surgery, urinary NT significantly increase. This raise was remarkably higher in women who develop urgency, who also presented a higher reduction in Qmax. These data suggest that obstruction may cause long-term changes in urinary NT, promoting the emergence of urgency. © 2014FiguresReferencesRelatedDetails Volume 191Issue 4SApril 2014Page: e884 Advertisement Copyright & Permissions© 2014MetricsAuthor Information Tiago Antunes-Lopes More articles by this author Daniel Costa More articles by this author Rui Pinto More articles by this author Sérgio Carvalho-Barros More articles by this author Carlos Martins-Silva More articles by this author Célia Duarte-Cruz More articles by this author Francisco Cruz More articles by this author Expand All Advertisement Advertisement PDF DownloadLoading ...

MP1-09 ASSOCIATION BETWEEN AGE-RELATED DETRUSOR UNDERACTIVITY AND DOWNREGULATION OF THE BIOLOGICAL CLOCK OF CONNEXIN 43-DERIVED GAP JUNCTIONS IN RAT BLADDERS

INTRODUCTION AND OBJECTIVES: Various diseases can cause altered afferent autonomic sensitivity of bladder, including diabetes, neurologic or vascular lesions, multiple sclerosis, recurrent infection, interstitial cystitis, pelvic pain syndrome. In current urologic clinical practice, to study autonomic and sensory bladder neuropathy, clinicians still rely on patientsi?/2 self-reported bladder sensation during the filling phase of urodynamics. We developed and tested the efficacy of an implantable bladder device which, when combined with the Neurometeri?/2, can be used to assess fiber specific afferent bladder sensation in the mouse. METHODS: We developed an implantable bladder device that applies selective nerve fiber stimuli (250 Hz for small myelinated Ad fibers and 5 Hz for unmyelinated C fibers) to the bladder mucosa in the mouse to determine bladder current perception threshold (CPT) values. We performed 4 experiments in 64 C57BL/6J mice to examine the effects of our device on voiding habits, assess the interobserver reliability of the sensory perception threshold and determine the effects of intravesical administration of resiniferatoxin and lidocaine on the sensory perception threshold. RESULTS: Sensory perception threshold values obtained by 2 blinded, independent observers were not different from each other (p1⁄40.74382). CPT values obtained at the 2 stimulation frequencies remained constant for at least 6 days after device implantation. A significant increase in CPT values after resiniferatoxin instillation was noted at a stimulus frequency of 5 Hz (p1⁄40.01347), whereas intravesical lidocaine led to an immediate increase in CPT at 250 (p1⁄40.0003) and 5 Hz (p1⁄40.0001). The implantation led to an early decreased voided volume and increased frequency of voids at 4 days, although these parameters returned to close normal after 6 days. The implanted bladder histology reveals defect created by device and mild inflammatory changes in next areas. CONCLUSIONS: Assessment of bladder afferent sensation with our newly developed device is feasible in mice. It provides CPTs that appear to be fiber-type selective for autonomic bladder afferent nerves.

MP1-03 HERPES SIMPLEX VIRUS (HSV) VECTOR-MEDIATED GENE DELIVERY OF PROTEIN PHOSPHATASE 1α REDUCES BLADDER OVERACTIVITY AND NOCICEPTION IN RATS

You have accessJournal of UrologyUrodynamics/Incontinence/Female Urology: Basic Research I1 Apr 2014MP1-03 HERPES SIMPLEX VIRUS (HSV) VECTOR-MEDIATED GENE DELIVERY OF PROTEIN PHOSPHATASE 1α REDUCES BLADDER OVERACTIVITY AND NOCICEPTION IN RATS Tsuyoshi Majima, Naoki Kawamorita, Hiroki Okada, Yasuhito Funahashi, Momokazu Gotoh, William Goins, Joseph Glorioso, Justus Cohen, Bonnie Hall, Asaff Harel, and Naoki Yoshimura Tsuyoshi MajimaTsuyoshi Majima More articles by this author , Naoki KawamoritaNaoki Kawamorita More articles by this author , Hiroki OkadaHiroki Okada More articles by this author , Yasuhito FunahashiYasuhito Funahashi More articles by this author , Momokazu GotohMomokazu Gotoh More articles by this author , William GoinsWilliam Goins More articles by this author , Joseph GloriosoJoseph Glorioso More articles by this author , Justus CohenJustus Cohen More articles by this author , Bonnie HallBonnie Hall More articles by this author , Asaff HarelAsaff Harel More articles by this author , and Naoki YoshimuraNaoki Yoshimura More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2014.02.101AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Increased afferent excitability has been proposed as an important pathophysiological basis of overactive bladder (OAB) and hypersensitive bladder disorders such as interstitial cystitis/bladder pain syndrome (IC/BPS). It has also been reported that transient receptor potential vanilloid-1 (TRPV1) receptors predominantly expressed in C-fiber afferent pathways greatly contribute to afferent sensitization in these disease conditions. We recently found Protein Phosphatase 1α (PP1α) as a modulator of TRPV1 receptor activity using an HSV vector cDNA library screen method to identify genes that inhibit TRPV1 activation, and showed that HSV vector-mediated PP1α expression led to the reduction in capsaicin-induced thermal hyperalgesia. Therefore, this led us to investigate the effect of HSV vectors-mediated gene delivery of PP1α on TRPV1-mediated bladder overactivity and pain-related behavior in rats. METHODS Replication-deficient HSV vectors encoding PP1α or green fluorescent protein (GFP) as control were injected into the bladder wall of adult female Sprague-Dawley rats. Cystometry (CMG) under urethane anesthesia was performed 1 week after viral injection to evaluate bladder overactivity induced by resiniferatoxin (RTX, a TRPV1 agonist). RTX-induced nociceptive behavior such as licking (lower abdominal licking) and freezing (motionless head-turning) was observed 2 weeks after viral injection. Using immunohistochemistry, GFP expression in L6/S1 DRG and the bladder as well as c-Fos positive cells in the L6 spinal cord dorsal horn were evaluated. RESULTS GFP expression was seen in L6/S1 DRG sections. In CMG, the PP1α group showed a significantly (p<0.05) smaller reduction (46.5±1.7%) in intercontraction intervals after RTX infusion than the GFP group (65.7±3.8%). The number of RTX-induced freezing behavior, which is correlated with bladder pain sensation, was significantly (p<0.001) decreased in PP1α vs. GFP groups. The number of c-Fos positive cells in the L6 spinal dorsal horn was significantly (p<0.01) smaller in PP1α vs. GFP rats (24±4 vs. 59±6 cells per section). CONCLUSIONS These results indicate that HSV vectors injected into the bladder wall are transported to lumbosacral DRGs that contain bladder afferent neurons, and that PP1α gene delivery in the bladder is effective in suppressing TRPV1-mediated bladder overactivity and pain behavior. Thus, HSV-mediated PP1α gene therapy could be effective for the treatment of OAB and/or hypersensitive bladder disorders such as IC/BPS. © 2014FiguresReferencesRelatedDetails Volume 191Issue 4SApril 2014Page: e1-e2 Peer Review Report Advertisement Copyright & Permissions© 2014MetricsAuthor Information Tsuyoshi Majima More articles by this author Naoki Kawamorita More articles by this author Hiroki Okada More articles by this author Yasuhito Funahashi More articles by this author Momokazu Gotoh More articles by this author William Goins More articles by this author Joseph Glorioso More articles by this author Justus Cohen More articles by this author Bonnie Hall More articles by this author Asaff Harel More articles by this author Naoki Yoshimura More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

The Muscle Stretch Reflex throughout the Menstrual Cycle

The significant sex disparity in sports-related knee injuries may be due to underlying differences in motor control. Although the development of sex-specific movement patterns is likely multifactorial, this study specifically focuses on the potential modulatory role of sex hormones. This study aimed to investigate the muscle stretch reflex (MSR) across the menstrual cycle. We hypothesized that the MSR would fluctuate throughout the menstrual cycle and that the lowest response would correspond with peak concentrations of estrogen. Nineteen healthy women age 18-35 yr participated in this study: 8 eumenorrheic women and 11 women taking oral contraceptives. Serum estradiol and progesterone concentrations, anterior knee laxity (AKL), and the MSR response of the quadriceps muscles were measured three times during the menstrual cycle. The MSR response of the rectus femoris (RF) varied significantly across the menstrual cycle in both groups. Specifically, the RF MSR response was 2.4 times lower during the periovulatory phase when compared with the luteal phase (P = 0.007). The same trend was seen in the vastus medialis, but this did not reach statistical significance (P = 0.070). The MSR response of the vastus lateralis did not change significantly across the menstrual cycle (P = 0.494). A mixed model comparison did not show an association between endogenous concentrations of estradiol and progesterone, exposure to hormonal contraceptives or AKL, and the MSR response for any muscle. Our results demonstrate that the RF MSR response varies throughout the menstrual cycle with the lowest response around the time of ovulation. Additional research is needed to clarify the exact relationship between sex hormones, AKL, and MSR response and to determine the specific origin of the change along the monosynaptic reflex arc.