Célia Duarte Cruz

Inhibition of ERK phosphorylation decreases nociceptive behaviour in monoarthritic rats.

&NA; In this study we investigated the role of the activation of the extracellular signal‐regulated kinases 1 and 2 (ERK) in chronic inflammatory articular nociception. Monoarthritis was induced in the left ankle of Wistar rats by injection of complete Freunds adjuvant (CFA). Movement of the inflamed joint increased ERK phosphorylation in neurones of the superficial and deep ipislateral dorsal horn laminae of L3‐L5 spinal cord segments. Spinal immunoreactivity to phosphoERK was more intense in animals in which the inflammation lasted longer, 7 days or more, than in rats with less time of inflammation. PhosphoERK levels were transient, since 2 h after ankle stimulation spinal immunoreaction had almost disappeared. PhosphoERK immunoreactivity was not induced by movement of ankles from non‐arthritic control animals, neither in monoarthritic rats in which the inflamed ankle was not stimulated. Intrathecal administration of PD 98059, an inhibitor of ERK phosphorylation, reduced nociceptive behaviour induced by the ankle bend test in monoarthritic rats. The anti‐nociceptive effect of PD 98059 was more prominent and in animals with short lasting (4 days) than in animals with longer (14 days) monoarthritis. Taken together, these findings suggest that ERK phosphorylation in spinal cord neurones plays an important role in chronic inflammatory articular pain and that its inhibition may provide significant anti‐nociception.

Neurotrophins as regulators of urinary bladder function

Increased voiding frequency and urgency are among the most prevalent storage lower urinary tract symptoms (LUTS), often diagnosed as part of overactive bladder syndrome (OAB). It has been suggested that these symptoms are caused by excessive sensory activation of the neural micturition circuit. It seems likely that sensory pathway remodelling is also responsible for pain perception upon bladder filling in patients with bladder pain syndrome (BPS). Neurotrophins—including nerve growth factor (NGF), brain-derived nerve factor (BDNF), neurotrophin-3 (NT-3) and neurotrophin-4 (NT-4)—represent master modulators of neural plasticity, both in peripheral and central nervous systems. Accumulating evidence points towards a role for neurotrophins in the control of neural sensory function during micturition and indicates their involvement in the emergence of OAB-related and BPS-related LUTS. Neurotrophins could potentially be used as urinary biomarkers to improve diagnostic accuracy for OAB and BPS and monitor therapy effectiveness. Proof-of-principle clinical evidence has confirmed that NGF is a potential target for treating human bladder overactivity.

The ERK 1 and 2 Pathway in the Nervous System: From Basic Aspects to Possible Clinical Applications in Pain and Visceral Dysfunction

The extracellular signal-regulated kinases 1 and 2 (ERK) cascade, member of the mitogen-activated protein kinases superfamily of signalling pathways, is one of the best characterized pathways as many protein interactions and phosphorylation events have been systematically studied. Traditionally, ERK are associated with the regulation of proliferation and differentiation as well as survival of various cell types. Their activity is controlled by phosphorylation on specific aminoacidic residues, which is induced by a variety of external cues, including growth-promoting factors. In the nervous system, ERK phosphorylation is induced by binding of neurotrophins to their specific tyrosine kinase receptors or by neuronal activity leading to glutamate release and binding to its ionotropic and metabotropic receptors. Some studies have provided evidence of its importance in neuroplastic events. In particular, ERK phosphorylation in the spinal cord was shown to be nociceptive-specific and its upregulation, occurring in cases of chronic inflammatory and neuropathic pain, seems to be of the utmost importance to behavioural changes observed in those conditions. In fact, experiments using specific inhibitors of ERK phosphorylation have proved that ERK directly contributes to allodynia and hyperalgesia caused by spinal cord injury or chronic pain. Additionally, spinal ERK phosphorylation regulates the micturition reflex in experimental models of bladder inflammation and chronic spinal cord transection. In this review we will address the main findings that suggest that ERK might be a future therapeutic target to treat pain and other complications arising from chronic pain or neuronal injury.

The activation of the ERK pathway contributes to the spinal c-fos expression observed after noxious bladder stimulation.

C-fos is an immediate-early gene whose expression in the spinal cord has been extensively used as a marker of peripheral noxious stimulation. The Fos protein accumulates in the nuclei of spinal neurons, reaching detectable levels 2u2009h after stimulation. The ERK pathway is an important signalling pathway in spinal cord neurons. ERK is activated upon phosphorylation on specific amino acid residues. Its activation in the spinal cord, following noxious stimulation, has been shown to contribute to the establishment and maintenance of long-term neuronal alterations associated with chronic pain. Phosphorylated ERK can target several cellular elements, including transcription factors, which indicates that ERK participates in the regulation of gene expression. The relation between ERK and c-fos is at present still unclear. Some in vitro studies have reached the conclusion that ERK contributes to c-fos regulation whereas others have provided evidence of ERK-independent c-fos expression. In fact, in the spinal cord the occurrence of c-fos expression in the absence of ERK phosphorylation has been reported. In this study we investigated in vivo the contribution of ERK to c-fos expression in the spinal cord. By inhibiting spinal ERK activation with intrathecal administration of PD98059, we verified that ERK phosphorylation does contribute to regulate c-fos expression upon noxious bladder stimulation.

Spinal ERK activation contributes to the regulation of bladder function in spinal cord injured rats

The extracellular signal-regulated kinase 1 and 2 (ERK) pathway, regulated by phosphorylation on specific amino acids, is emerging as an important signaling cascade in neurones, transducing sensory input into cellular responses. In the mammalian nervous system, the ERK pathway has been found to mediate plasticity events. Particularly, in the spinal cord, ERK play an important role in allodynia and hyperalgesia. Recently, it was demonstrated that ERK activation is upregulated in the spinal cord of rats with chronic bladder inflammation and contributes to bladder overactivity. Thus, in this study we sought to assess the involvement of ERK in micturition reflexes associated to spinal cord injury (SCI) in the rat. Bladder function in chronic SCI rats was altered compared to spinal intact rats. PhosphoERK levels were upregulated in the L6 spinal cord segment, particularly after saline infusion for 2 h. The increase in spinal ERK phosphorylation was specifically restricted to L6 spinal segment. No variation in the levels of total ERK protein was observed. Intrathecal administration of PD98059, a specific inhibitor of ERK phosphorylation, reduced the frequency and amplitude of bladder contractions in SCI animals but not in spinal intact ones. Overall, our results demonstrate increased activation of the ERK pathway in the spinal cord from SCI rats, restricted to spinal segments that receive sensory input arising from the bladder. Since the use of PD98059 reduced the frequency and amplitude of bladder contractions, ERK inhibitors may provide a new therapeutic approach to the treatment of bladder overactivity after spinal injuries.

Biomarkers in lower urinary tract symptoms/overactive bladder: a critical overview.

Purpose of review Biomarkers constitute objectively measurable characteristics that can be evaluated as indicators of physiological and pathogenic processes and might be used as diagnostic, prognostic or predictive tools in clinical care. This review examines the availability of biomarkers to treat the dynamic and complex symptoms of overactive bladder (OAB). Recent findings OAB biomarkers may contribute to reveal the origin of storage symptoms in otherwise healthy individuals. The research encompassing the changes that occur in the bladder or in the peripheral (and central) nervous system might be determined through blood or urinary molecules (neurotrophins, ATP, prostaglandins, C-reactive protein and cytokines) or the measurement of events occurring in the bladder wall (bladder wall or detrusor wall thickness, oxyhemoglobin and deoxyhemoglobin concentration). These biomarkers might contribute to a better understanding of the pathophysiologic mechanisms underlying OAB. Summary The word biomarker to name all the parameters described above, from bladder wall thickness to urinary molecules, has been introduced to call the attention to a field wherein objective noninvasive parameters were nonexistent. OAB treatment based on a biomarker, in comparison to the treatment based on a diagnosis made from a careful history and exclusion of urinary tract infection, is not supported by current literature.

Neurotrophins in the lower urinary tract: becoming of age.

The lower urinary tract (LUT) comprises a storage unit, the urinary bladder, and an outlet, the urethra. The coordination between the two structures is tightly controlled by the nervous system and, therefore, LUT function is highly susceptible to injuries to the neuronal pathways involved in micturition control. These injuries may include lesions to the spinal cord or to nerve fibres and result in micturition dysfunction. A common trait of micturition pathologies, irrespective of its origin, is an upregulation in synthesis and secretion of neurotrophins, most notably Nerve Growth Factor (NGF) and Brain Derived Neurotrophic Factor (BDNF). These neurotrophins are produced by neuronal and non-neuronal cells and exert their effects upon binding to their high-affinity receptors abundantly expressed in the neuronal circuits regulating LUT function. In addition, NGF and BDNF are present in detectable amounts in the urine of patients suffering from various LUT pathologies, suggesting that analysis of urinary NGF and BDNF may serve as likely biomarkers to be studied in tandem with other factors when diagnosing patients. Studies with experimental models of bladder dysfunction using antagonists of NGF and BDNF receptors as well as scavenging agents suggest that those NTs may be key elements in the pathophysiology of bladder dysfunctions. In addition, available data indicates that NGF and BDNF might constitute future targets for designing new drugs for better treatment of bladder dysfunction.

MP42-06 EXPRESSION AND FUNCTION OF SEROTONIN PARANEURONAL CELLS IN THE URETHRAL EPITHELIUM OF HUMAN AND RODENTS

persists following the resolution of colitis, and that linaclotide, an FDA approved guanylate cyclase-C (GC-C) agonist, is able to attenuate these changes [3]. We hypothesise that these CCH-induced changes are the result of altered sensitivity of afferents both within the colon and bladder wall and within the dorsal root ganglion (DRG), and that oral linaclotide administration may act to reduce this hypersensitivity. METHODS: We investigated healthy C57BL/6J mice and mice with CCH, 28 days after intra-colonic TNBS administration. CCH mice were randomly assigned to either chronic linaclotide (3mg/kg/day) or placebo (water) administration, consisting of a once daily oral gavage for 2 weeks prior to experimentation. Ex-vivo electrophysiological recordings determined bladder afferent and contractile sensitivity to abMe-ATP (30mM), carbachol (1mM), and capsaicin (10mM) as well as whole cell patch clamp of retrogradely traced bladder DRG neurons in all four groups. RESULTS: Bladder DRG from mice with CCH display hyperexcitability, with a significant reduction in rheobase compared to controls (p 0.01). CCH mice also display significantly enhanced afferent responses to abMe-ATP (p 0.001), carbachol (p 0.001), and capsaicin (p 0.001). CCH mice treated with linaclotide display attenuated DRG hyperexcitability and normalised afferent responses to agonists (p 0.01) compared to placebo (p 0.01). CONCLUSIONS: Mice with CCH also display increased bladder afferent excitability within both the DRG and bladder wall, indicating cross-organ sensitisation. Chronic oral administration of linaclotide, a locally acting GC-C agonist that inhibits colonic nociceptors, reverses these colitis-induced changes in bladder afferent sensitivity. Common sensory pathways may allow agents that reduce abdominal pain to improve urological symptoms. 1. Lamb, K., et al. AJPGI, 2006. 2. Ustinova et al, Neurourology and Urodynamics, 2010. 3. Grundy, L., et al., MP28-06. The Journal of Urology, 2016.

Biomarkers in the Overactive Bladder Syndrome

Overactive bladder (OAB) is a symptomatic complex affecting both men and women. The overall incidence is above 10% but may exceed 40% in the elderly population (Irwin et al., 2006; Irwin et al., 2009; Sexton et al., 2009a). OAB is defined by the International Continence Society (ICS) as a clinical syndrome characterized by urinary urgency, with or without incontinence, usually with frequency and nocturia (Abrams et al., 2002, 2003; Hashim & Abrams, 2007). Urgency, which is a storage symptom defined as a sudden compelling desire to pass urine difficult to defer, is the hallmark symptom as it is the only one that must be present in order to establish the diagnosis of OAB (Abrams et al., 2002, 2003). Several comorbidities are very common among OAB patients, including depression, insomnia and fractures (Coyne et al., 2008; Sexton et al., 2009a; Sexton et al., 2009b). The economic costs of OAB, associated with medical consultations, therapy and diminished productivity at work, may reach billions of dollars (Irwin et al., 2009) and will certainly increase with the demographic shift of an ageing population.

An Update on Neurotrophins

Neurotrophins are important proteins that regulate survival, development and function of neurons. They are produced by a variety of cells and exert their effects upon binding to specific tyrosine kinase receptors. The first neurotrophin to be identified was Nerve Growth Factor (NGF). From the pioneer studies performed by Rita-Levi Montalcini, awarded with the Nobel Prize, other neurotrophins have been identified and this family of neurotrophic factors now includes Brain Derived Neurotrophic Factor (BDNF), Neurotrophin 3 and Neurotrophin 4. Their function as pro-survival proteins has been consolidated but subsequent studies have demonstrated that neurotrophins may also play a role in some pathological conditions. This is the focus of this special issue of Current Neuropharmacology. n nThe paper by Allen et al., addresses the involvement of neurotrophins, NGF and BDNF in particular, in Alzheimer’s disease and the possibility of using neurotrophin-based therapies. n nFrias et al., summarize studies that support the involvement of neurotrophins in the lower urinary tract (LUT) dysfunction and suggest the use of urinary neurotrophins as biomarkers of LUT pathologies. n nThe paper by Neto et al., addresses the involvement of neurotrophins in depression, a poorly understood pathology with an increasing prevalence in Western countries. n nThe paper by Siniscalco et al., addresses the involvement of neurotrophins in neuropathic pain, a complex pathology difficult to tackle with, and its importance as targets for neuropathic pain treatment. n nTeng et al., summarize the clinical uses of stem cell-based administration of neurotrophins for the treatment of spinal cord injury. n nThis issue is published in memory of David Dawbarn, who passed away unexpectedly in January 2010. David dedicated most of his work at the Bristol University to understanding the importance of neurotrophic factors in Alzheimer’s disease, unraveling the molecular interactions between neurotrophins and their high-affinity receptors. His research was a key factor to the design of new therapies for the treatment of Alzheimer’s disease, pain and asthma. He is greatly missed by the scientific community, friends and family.