Neuropathic and inflammatory antinociceptive effects and electrocortical changes produced by Salvia divinorum in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Salvia divinorum is a medicinal plant traditionally used in hallucinogenic ethnopharmacological practices and for its analgesic and antinflammatory properties. Its active compounds include diterpenes known as salvinorins which act as potent κ opioid receptor agonists. AIM OF THE STUDY: Given its effects in acute animal models of pain, as well as its antinflammatory attributes, we decided to investigate the analgesic effects of an SD extract in neuropathic (sciatic loose nerve ligature) and inflammatory (intra plantar carrageenan) pain models in rats. We also determined in this study the electrocorticographic changes to correlate similar hallucinogenic state and behavior as those produced in humans. MATERIAL AND METHODS: Mechanical and thermonociceptive responses, plantar test and von Frey assay, respectively, were measured in adult Wistar rats 30min, 3h and 24h after the intraperitoneal administration of saline or an hydroponic SD extract. We also evaluated carbamazepine and celecoxib, as gold reference drugs, to compare its antinociceptive effects. RESULTS: Our results showed that administration of SD extract induced antialgesic effects in both neuropathic and inflammatory pain models. All those effects were blocked by nor-binaltorphimine (a Kappa opioid receptor antagonist). Moreover, it was observed an increase of the anterior power spectral density and a decrease in the posterior region as electrocorticographic changes. CONCLUSION: The present investigation give evidence that SD is capable to reduce algesic response associated to neuropathic and inflammatory nociception. This study support therapeutic alternatives for a disabling health problem due to the long term pain with high impact on population and personal and social implications.

A D2-like receptor family agonist produces analgesia in mechanonociception but not in thermonociception at the spinal cord level in rats.

The administration of dopaminergic drugs produces analgesia in individuals experiencing different types of pain. Analgesia induced by these drugs at the spinal cord level is mediated by D2-like agonists, which specifically inhibit the detection of nociceptive stimuli by sensory afferents. The extent of the analgesia provided by spinal dopamine agonists remains controversial, and the cellular mechanism of this analgesic process is poorly understood. The objective of this study was to evaluate the analgesic effect of quinpirole, a D2-like agonist, based on two nociceptive tests and at various doses that were selected to specifically activate dopamine receptors. We found that intrathecal quinpirole administration produces analgesia of mechanical but not thermal nociception and that the analgesic effect of quinpirole is reversed by a mix of D2, D3, and D4 receptor-specific antagonists, suggesting that the activation of all D2-like receptors is involved in the analgesia produced by intrathecal quinpirole. The differential effect on thermal and mechanical nociception was also tested upon the activation of µ-opioid receptors. As reported previously, low doses of the µ-opioid receptor agonist DAMGO produced analgesia of only thermonociception. This evidence shows that a D2-like receptor agonist administered at the spinal cord level produces analgesia specific to mechanonociception but not thermonociception.

Inhibition of peripheral nociceptors by aminoglycosides produces analgesia in inflammatory pain models in the rat.

Aminoglycosides (AGs) modulate nociceptors and ionic channels expressed in sensory neurons. The AG applied in situ could be useful to alleviate hyperalgesia in animal models of inflammatory pain. We tested streptomycin (ST) and neomycin (NEO) as analgesic agents applied in situ in rat paw inflammation caused by formalin or carrageenan administration. The action of ST and NEO on the action potential discharge produced by acidic stimuli in isolated dorsal root ganglion neurons was also studied in current-clamp recordings. In the formalin test, ST and NEO significantly reduced the nociceptive behaviour. ST reduced the N-(4-methyl-2-quinazolinyl)-guanidine (GMQ)-induced nociceptive behaviour, and NEO diminished the hyperalgesia to thermonociception and mechanonociception produced by CAR. In the current-clamp experiments, ST and NEO reduced the generation of action potentials when an acidic solution was applied. We conclude that ST and NEO produce analgesia to inflammatory pain, an effect that is due in part to the inhibition of ASIC activation in sensory neurons.

Inflammatory nociception responses do not vary with age, but diminish with the pain history.

Some of the relevant factors that must be considered when dealing with old age include its growing numbers in the general population and pain contention in this age group. In this sense, it is important to study whether antinociceptive responses change with age. To elucidate this point, persistent pain in animals is the preferred model. In addition, the response to inflammatory pain in the same individual must be explored along its lifetime. Male Wistar rats were infiltrated with carrageenan (50 µl intraplantar) and tested 3 h and 24 h after injection using thermal (plantar test) and mechanociceptive tests (von Frey). The rats were divided into the following groups: (a) young rats infiltrated for the first time at 12 weeks of age and re-infiltrated at 15 and 17 weeks; (b) adult rats infiltrated for the first time at 28 weeks of age and re-infiltrated at 44 and 56 weeks; and (c) old rats infiltrated for the first time at 56 weeks of age and re-infiltrated at 72 weeks. The rats tested for the first time at 12 and 56 weeks of age showed hyperalgesia due to carrageenan infiltration at 3 h and 24 h after injection. This result showed that old rats maintain the same antialgesic response due to inflammation. However, when the injection was repeated in the three age groups, the latency to the thermal and mechanociceptive responses at 3 h is increased when compared to animals exposed for the first time to inflammation. The response to thermal and mechanociception in old rats is the same as in young animals as long as the nociceptive stimulus is not repeated. The repetition of the stimulus produces changes compatible with desensitization of the response and evidences the significance of algesic stimulus repetition in the same individual rather than the age of the individual.

Taurine enhances antinociception produced by a COX-2 inhibitor in an inflammatory pain model.

The temporal activation of the sensory systems, especially in pain, determines intermediate states that define the future of the response to sensory stimulation. In this work, we interfere pharmacologically with those states that produce peripheral and central sensitisation after an acute inflammatory process, inhibiting at the periphery the COX-2 with celecoxib and using taurine (glycine A receptor agonist) for central pain relief. We tested the paw withdrawal reflex latencies to thermo- and mechanonociception after the induction of an acute inflammatory process with carrageenan. Celecoxib at low doses [0.13 and 1.3 mg/kg, intraperitoneal (i.p.)] in combination with taurine (300 mg/kg, i.p.) produces a decrease of the nociceptive response in thermo- and mechanonociception, as compared with the effect of both drugs alone. We propose that the enhancement of the analgesic effect of celecoxib in combination with taurine could be due the simultaneous action of these drugs at both, peripheral and central levels.

Procesamiento central del dolor neuropático: una aproximación integrativa / Central processing of neuropathic pain: an integrative approach

The term pain matrix refers to the structures and pathways in the central nervous system that play a role in pain processing and integration. For the last several years, our group has been studying the mechanisms that are involved in the establishment of long-term pain. Our research focus has been the study of the different nuclei and corticolimbic pathways that are involved in the affective-cognitive component of pain. In addition, we have also explored painful processes and memory. The pain matrix is constituted by the ventral tegmental area (VTA), anterior cingulate cortex (ACC), and insular cortex, among others. VTA is a predominantly dopaminergic area and has projections to ACC and the insular cortex. Stimulation of this region can reduce nociception, whereas its lesion has the opposite effect. In the ACC, it has been studied how excitatory aminoacids, such as glutamate, increase nociception while inhibitory ones decrease it. Moreover, this cortex is associated with mechanisms of pain memory. In this sense, we have seen that blocking cholinergic receptors diminishes the acquisition of pain-related memories. Nociceptive stimuli increase the expression of inhibitory muscarinic M2 receptors. In relation with insular cortex, the focus of study has been on the dopaminergic system. We have found that blocking dopaminergic D2 receptors significantly reduces neuropathic nociception. In response to an inflammatory process there is a decrease in the extracellular levels of dopamine and in the expression of mRNA for excitatory dopamine D1 receptors, while there is an increase in mRNA expression for inhibitory D2 receptors. Despite current progress in this research area, more studies are needed in order to integrate the relationship among the different neurotransmission systems. This will contribute to the proposal of novel therapeutic alternatives to the conventional treatments for pain.

El término "matriz del dolor" se refiriere a todas las estructuras y vías del Sistema Nervioso Central relacionadas con la integración del dolor. Nuestro grupo estudia desde hace varios años los principales mecanismos involucrados en el desarrollo del dolor a largo plazo. Nos hemos enfocado en el estudio de diferentes núcleos y vías cortico-límbicas que están relacionadas con la parte afectiva-cognitiva, así como en la memoria de los procesos dolorosos. Dentro de estos núcleos se encuentra el área tegmental ventral (ATV), la corteza anterior del cíngulo (CAC) y la corteza insular. El ATV es una estructura principalmente dopaminérgica con proyecciones a la CAC y a la corteza insular. Como se verá más adelante, estimular este núcleo disminuye la nocicepción, mientras que el lesionarlo, la aumenta. En la CAC se ha estudiado cómo aminoácidos excitadores como el glutamato aumentan la nocicepción y cómo, por el contrario, los aminoácidos inhibitorios como la taurina, la disminuyen. Además esta corteza está relacionada con mecanismos de memoria dolorosa. Hemos visto que el bloqueo de receptores colinérgicos disminuye la adquisición de la memoria relacionada al dolor. Además, un estímulo nociceptivo aumenta la expresión de los receptores muscarínicos inhibitorios M2. En el caso de la corteza insular, se ha estudiado principalmente el papel del sistema dopaminérgico. Hemos encontrado que el bloqueo de receptores dopaminérgicos D2 disminuye de manera significativa la nocicepción neuropática. Encontramos también que los niveles extracelulares de dopamina en esta región disminuyen a consecuencia de un proceso inflamatorio, además de que disminuye la expresión del RNAm de los receptores excitadores D1 y aumenta la de los receptores inhibidores D2. A pesar del avance que se ha obtenido en esta área de investigación, se necesitan más estudios para integrar la relación entre los diferentes sistemas de neurotransmisión y poder proponer alternativas a los tratamientos convencionales para las diferentes patologías que cursan con una experiencia dolorosa.

Nras loss induces metastatic conversion of Rb1-deficient neuroendocrine thyroid tumor.

Mutations in the gene encoding the retinoblastoma tumor suppressor predispose humans and mice to tumor development. Here we have assessed the effect of Nras loss on tumor development in Rb1 heterozygous mice. Loss of one or two Nras alleles is shown to significantly reduce the severity of pituitary tumors arising in Rb1(+/-) animals by enhancing their differentiation. By contrast, C-cell thyroid adenomas occurring in Rb1(+/-) mice progress to metastatic medullary carcinomas after loss of Nras. In Rb1(+/-)Nras(+/-) animals, distant medullary thyroid carcinoma metastases are associated with loss of the remaining wild-type Nras allele. Loss of Nras in Rb1-deficient C cells results in elevated Ras homolog family A (RhoA) activity, and this is causally linked to the invasiveness and metastatic behavior of these cells. These findings suggest that the loss of the proto-oncogene Nras in certain cellular contexts can promote malignant tumor progression.

Genetic interaction between Rb and K-ras in the control of differentiation and tumor suppression.

Although the retinoblastoma protein (pRb) has been implicated in the processes of cellular differentiation, there is no compelling genetic or in vivo evidence that such activities contribute to pRb-mediated tumor suppression. Motivated by cell culture studies suggesting that Ras is a downstream effector of pRb in the control of differentiation, we have examined the tumor and developmental phenotypes of Rb and K-ras double-knockout mice. We find that heterozygosity for K-ras (i) rescued a unique subset of developmental defects that characterize Rb-deficient embryos by affecting differentiation but not proliferation and (ii) significantly enhanced the degree of differentiation of pituitary adenocarcinomas arising in Rb heterozygotes, leading to their prolonged survival. These observations suggest that Rb and K-ras function together in vivo, in the contexts of both embryonic and tumor development, and that the ability to affect differentiation is a major facet of the tumor suppressor function of pRb.

A mechanism of cyclin D1 action encoded in the patterns of gene expression in human cancer.

Here we describe how patterns of gene expression in human tumors have been deconvoluted to reveal a mechanism of action for the cyclin D1 oncogene. Computational analysis of the expression patterns of thousands of genes across hundreds of tumor specimens suggested that a transcription factor, C/EBPbeta/Nf-Il6, participates in the consequences of cyclin D1 overexpression. Functional analyses confirmed the involvement of C/EBPbeta in the regulation of genes affected by cyclin D1 and established this protein as an indispensable effector of a potentially important facet of cyclin D1 biology. This work demonstrates that tumor gene expression databases can be used to study the function of a human oncogene in situ.

Rb and N-ras function together to control differentiation in the mouse.

The product of the retinoblastoma tumor suppressor gene (Rb) can control cell proliferation and promote differentiation. Murine embryos nullizygous for Rb die midgestation with defects in cell cycle regulation, control of apoptosis, and terminal differentiation of several tissues, including skeletal muscle, nervous system, and lens. Previous cell culture-based experiments have suggested that the retinoblastoma protein (pRb) and Ras operate in a common pathway to control cellular differentiation. Here we have tested the hypothesis that the proto-oncogene N-ras participates in Rb-dependent regulation of differentiation by generating and characterizing murine embryos deficient in both N-ras and Rb. We show that deletion of N-ras rescues a unique subset of the developmental defects associated with nullizygosity of Rb, resulting in a significant extension of life span. Rb(-/-); N-ras(-/-) skeletal muscle has normal fiber density, myotube length and thickness, in contrast to Rb-deficient embryos. Additionally, Rb(-/-); N-ras(-/-) muscle shows a restoration in the expression of the late muscle-specific gene MCK, and this correlates with a significant potentiation of MyoD transcriptional activity in Rb(-/-); N-ras(-/-), compared to Rb(-/-) myoblasts in culture. The improved differentiation of skeletal muscle in Rb(-/-); N-ras(-/-) embryos occurs despite evidence of deregulated proliferation and apoptosis, as seen in Rb-deficient animals. Our findings suggest that the control of differentiation and proliferation by Rb are genetically separable.