Anxiolytic-like effects induced by blockade of transient receptor potential vanilloid type 1 (TRPV1) channels in the medial prefrontal cortex of rats

The endocannabinoid anandamide, in addition to activating cannabinoid type 1 receptors (CB1), may act as an agonist at transient receptor potential vanilloid type 1 (TRPV1) channels. In the periaqueductal gray, CB1 activation inhibits, whereas TRPV1 increases, anxiety-like behavior. In the medial prefrontal cortex (mPFC), another brain region related to defensive responses, CB1 activation induces anxiolytic-like effects. However, a possible involvement of TRPV1 is still unclear. In the present study, we tested the hypothesis that TRPV1 channel contributes to the modulation of anxiety-like behavior in the mPFC. Male Wistar rats (n = 5-7 per group) received microinjections of the TRPV1 antagonist capsazepine (1-60 nmol) in the ventral portion of the mPFC and were exposed to the elevated plus maze (EPM) or to the Vogel conflict test. Capsazepine increased exploration of open arms in the EPM as well as the number of punished licks in the Vogel conflict test, suggesting anxiolytic-like effects. No changes in the number of entries into the enclosed arms were observed in the EPM, indicating that there were no changes in motor activity. Moreover, capsazepine did not interfere with water consumption or nociceptive threshold, discarding potential confounding factors for the Vogel conflict test. These data suggest that TRPV1 in the ventral mPFC tonically inhibits anxiety-like behavior. TRPV1 could facilitate defensive responses opposing, therefore, the anxiolytic-like effects reported after local activation of CB1 receptors.

Neuromuscular activity of BaTX, a presynaptic basic PLA(2) isolated from Bothrops alternatus snake venom

We have previously isolated a Lys49 phospholipase A(2) homolog (BaTX) from Bothrops alternatus snake venom using a combination of molecular exclusion chromatography and reverse phase HPLC and shown its ability to cause neuromuscular blockade. In this work, we describe a one-step procedure for the purification of this toxin and provide further details of its neuromuscular activity. The toxin was purified by reverse phase HPLC and its purity and molecular mass were confirmed by SIDS-PAGE, MALDI-TOF mass spectrometry, amino acid analysis and N-terminal sequencing. BaTX (0.007-1.4 mu M) produced time-dependent, irreversible neuromuscular blockade in isolated mouse phrenic nerve-diaphragm and chick biventer cervicis preparations (time to 50% blockade with 0.35 mu M toxin: 58 +/- 4 and 24 +/- 1 min, respectively; n = 3-8; mean +/- S.E.) without significantly affecting the response to direct muscle stimulation. In chick preparations, contractures to exogenous acetylcholine (55 and 110 mu M) or KCl (13.4 mM) were unaltered after complete blockade by all toxin concentrations. These results, which strongly suggested a presynaptic mechanism of action for this toxin, were reinforced by (1) the inability of BaTX to interfere with the carbachol-induced depolarization of the resting membrane, (2) a significant decrease in the frequency and amplitude of miniature end-plate potentials, and (3) a significant reduction (59 +/- 4%, n=12) in the quantal content of the end-plate potentials after a 60 min incubation with the toxin (1.4 mu M). In addition, a decrease in the organ bath temperature from 37 degrees C to 24 degrees C and/or the replacement of calcium with strontium prevented the neuromuscular blockade, indicating a temperature-dependent effect possibly mediated by enzymatic activity. (C) 2009 Elsevier Inc. All rights reserved.

Methods for exploring the morpho-functional relations of the aortic depressor nerve in experimental diabetes

The present study investigated morpho-functional relations of the aortic depressor nerve (ADN) 5, 15 and 120 days after the onset of streptozotocin-induced diabetes in rats. Time control animals received vehicle. Under pentobarbital anesthesia, ADN activity was recorded simultaneously with arterial pressure. After the recordings, nerves were prepared for light microscopy study and morphometry. ADN function was accessed by means of pressure-nerve activity curve (fitted by sigmoidal regression) and cross-spectral analysis between mean arterial pressure (MAP) and ADN activity. The relation between morphological (myelinated fibers number and density, total myelin area, total fiber area and percentage of occupancy) and functional (gain, signal/noise relation, frequency) parameters were accessed by linear regression analysis and correlation coefficient calculations. Functional parameters obtained by means of the sigmoidal regression curve as well as by cross-spectral analysis were similar in diabetic and control rats. Morphometric parameters of the ADN were similar between groups 5 days after the onset of diabetes. Average myelin area and myelinated fiber area were significantly smaller on diabetic rats 15 and 120 days after the onset of diabetes, being the myelinated fiber and respective axons area and diameter also smaller on 120 days group. Nevertheless, G ratio (ratio between axon and fiber diameter) was nearly 0.6 and not different between groups or experimental times. No significant relationship between morphological and functional parameters was detected in all experimental groups. The present study suggests that ADN diabetic neuropathy was time-dependent, with damage to myelinated fibers to be the primary event, not evidenced by physiological methods. (C) 2010 Elsevier B.V. All rights reserved.

Intermittent hypoxia-induced sensitization of central chemoreceptors contributes to sympathetic nerve activity during late expiration in rats

Molkov YI, Zoccal DB, Moraes DJ, Paton JF, Machado BH, Rybak IA. Intermittent hypoxia-induced sensitization of central chemoreceptors contributes to sympathetic nerve activity during late expiration in rats. J Neurophysiol 105: 3080-3091, 2011. First published April 6, 2011; doi:10.1152/jn.00070.2011.-Hypertension elicited by chronic intermittent hypoxia (CIH) is associated with elevated activity of the thoracic sympathetic nerve (tSN) that exhibits an enhanced respiratory modulation reflecting a strengthened interaction between respiratory and sympathetic networks within the brain stem. Expiration is a passive process except for special metabolic conditions such as hypercapnia, when it becomes active through phasic excitation of abdominal motor nerves (AbN) in late expiration. An increase in CO(2) evokes late-expiratory (late-E) discharges phase-locked to phrenic bursts with the frequency increasing quantally as hypercapnia increases. In rats exposed to CIH, the late-E discharges synchronized in AbN and tSN emerge in normocapnia. To elucidate the possible neural mechanisms underlying these phenomena, we extended our computational model of the brain stem respiratory network by incorporating a population of presympathetic neurons in the rostral ventrolateral medulla that received inputs from the pons, medullary respiratory compartments, and retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG). Our simulations proposed that CIH conditioning increases the CO(2) sensitivity of RTN/pFRG neurons, causing a reduction in both the CO(2) threshold for emerging the late-E activity in AbN and tSN and the hypocapnic threshold for apnea. Using the in situ rat preparation, we have confirmed that CIH-conditioned rats under normal conditions exhibit synchronized late-E discharges in AbN and tSN similar to those observed in control rats during hypercapnia. Moreover, the hypocapnic threshold for apnea was significantly lowered in CIH-conditioned rats relative to that in control rats. We conclude that CIH may sensitize central chemoreception and that this significantly contributes to the neural impetus for generation of sympathetic activity and hypertension.

Increased cardiac sympathetic drive and reduced vagal modulation following endothelin receptor antagonism in healthy conscious rats

1. The present study evaluated changes in autonomic control of the cardiovascular system in conscious rats following blockade of endothelin (ET) receptors with bosentan. 2. Rats were treated with bosentan or vehicle (5% gum arabic) for 7 days by gavage. 3. Baseline heart rate (HR) was higher in the bosentan-treated group compared with the control group (418 +/- 5 vs 357 +/- 4 b.p.m., respectively; P < 0.001). This baseline tachycardia was associated with a lower baroreflex sensitivity of the bradycardiac and tachycardiac responses in the bosentan-treated group compared with the control group. Sequential blockade of the parasympathetic and sympathetic autonomic nervous system with methylatropine and propranolol showed a higher intrinsic HR in the bosentan-treated group compared with the control group (411 +/- 5 vs 381 +/- 4 b.p.m., respectively; P < 0.05). This was accompanied by a higher cardiac sympathetic tone (31 +/- 1 vs 13 +/- 1%, respectively; P < 0.01) and a lower vagal parasympathetic tone (69 +/- 2 vs 87 +/- 2%, respectively; P < 0.01) in the bosentan-treated group compared with the control group. Variance and high-frequency oscillations of pulse interval (PI) variability in absolute and normalized units were lower in the bosentan-treated group than in the control group. Conversely, low-frequency (LF) oscillations of PI variability in absolute and normalized units, as well as variance and LF oscillations of systolic arterial pressure variability, were greater in the bosentan-treated group than the control group. 4. Overall, the data indicate an increased cardiac sympathetic drive, as well as lower vagal parasympathetic activity and baroreflex sensitivity, in conscious rats after chronic blockade of ET receptors with bosentan.

Morphology and morphometry of the vagus nerve in male and female spontaneously hypertensive rats

The vagus nerve is an important component of the efferent arm of the baroreflex. Blood pressure levels as well as baroreflex control of circulation are significantly different in male and female spontaneously hypertensive rats (SHR). We proposed to investigate the morphometric differences between genders using the vagus nerve of SHR. Adult animals (20 weeks old) were anesthetized and had their arterial pressure (AP) and heart rate (HR) recorded by a computerized system. The rats were then systemically perfused with a fixative solution and had their cervical vagi nerves prepared for light microscopy. Proximal and distal segments of the left and right vagi nerves were evaluated for morphometric parameters including fascicle area and diameter, myelinated fiber number, density, area and diameter. Comparisons were made between sides and segments on the same gender as well as between genders. Differences were considered significant when p<0.05. Male SHR had significantly higher AP and HR. Morphometric data showed no differences between the same levels of both sides and between segments on the same side for male and female rats. In addition, no significant morphometric differences were observed when genders were compared. This is the first description of vagus nerve morphometry in SHR indicating that gender differences in AP and HR cannot be attributed to dissimilarities in vagal innervation of the heart. These data provide a morphological basis for further studies involving functional investigations of the efferent arm of the baroreflex in hypertension. (C) 2007 Elsevier B.V. All rights reserved.

Toothache referred from auriculotemporal neuralgia: case report

P>Aim To present a 52-year-old male patient who complained of intense pain of short duration in the region of the left external ear and in the ipsilateral maxillary second molar that was relieved by blockade of the auriculotemporal nerve in the infratemporal fossa. Summary Extra- and intraoral physical examination revealed a trigger point that reproduced the symptoms upon finger pressure in the ipsilateral auriculotemporal nerve and in the outer auricular pavilion. The patient`s medical history was unremarkable. The maxillary left second molar tooth was not responsive to pulp sensitivity testing and there was no pain upon percussion or palpation of the buccal sulcus. Periapical radiographs revealed a satisfactory root filling in the maxillary left second molar. On the basis of the clinical signs and symptoms, the auriculotemporal was blocked with 0.5 mL 2% lidocaine and 0.5 mL of a suspension containing dexamethasone acetate (8 mg mL(-1)) and dexamethasone disodium sulfate (2 mg mL(-1)), with full remission of pain 6 months later. The diagnosis was auriculotemporal neuralgia. Key learning point Auriculotemporal neuralgia should be considered as a possible cause of nonodontogenic toothache and thus included in the differential diagnoses. The blockade of the auriculotemporal nerve in the infratemporal fossa is diagnostic and therapeutic. It can be achieved with a solution of lidocaine and dexamethasone.

Dorsal cutaneous branch of the ulnar nerve: a light and electron microscopy histometric study

This study describes the normal morphology and morphometry of the dorsal cutaneous branch of the ulnar nerve (DCBU) in humans. Fourteen nerves of eight donors were prepared by conventional techniques for paraffin and epoxy resin embedding. Semiautomatic morphometric analysis was performed by means of specific computer software. Histograms of the myelinated and unmyelinated fiber population and the G-ratio distribution of fibers were plotted. Myelinated fiber density per nerve varied from 5,910 to 10,166 fibers/mm(2), with an average of 8,170 +/- 393 fibers/mm(2). The distribution was bimodal with peaks at 4.0 and 9.5 mu m. Unmyelinated fiber density per nerve varied from 50,985 to 127,108, with an average of 78,474 +/- 6, 610 fibers/mm(2), with a unimodal distribution displaying a peak at 0.8 mu m. This study thus adds information about the fascicles and myelinated and unmyelinated fibers of DCBU nerves in normal people, which may be useful in further studies concerning ulnar nerve neuropathies, mainly leprosy neuropathy.

Nitric oxide synthesis blockade reduced the baroreflex sensitivity in trained rats

Objective: The present study has investigated the effect of blockade of nitric oxide synthesis on cardiovascular autonomic adaptations induced by aerobic physical training using different approaches: 1) double blockade with methylatropine and propranolol; 2) systolic arterial pressure (SAP) and heart rate variability (HRV) by means of spectral analysis; and 3) baroreflex sensitivity. Methods: Male Wistar rats were divided into four groups: sedentary rats (SR); sedentary rats treated with N(omega)-nitro-L-arginine methyl ester (L-NAME) for one week (SRL); rats trained for eight weeks (TR); and rats trained for eight weeks and treated with L-NAME in the last week (TRL). Results: Hypertension and tachycardia were observed in SRL group. Previous physical training attenuated the hypertension in L-NAME-treated rats. Bradycardia was seen in TR and TRL groups, although such a condition was more prominent in the latter. All trained rats had lower intrinsic heart rates. Pharmacological evaluation of cardiac autonomic tonus showed sympathetic predominance in SRL group, differently than other groups. Spectral analysis of HRV showed smaller low frequency oscillations (LF: 0.2-0.75 Hz) in SRL group compared to other groups. Rats treated with L-NAME presented greater LF oscillations in the SAP compared to non-treated rats, but oscillations were found to be smaller in TRL group. Nitric oxide synthesis inhibition with L-NAME reduced the baroreflex sensitivity in sedentary and trained animals. Conclusion: Our results showed that nitric oxide synthesis blockade impaired the cardiovascular autonomic adaptations induced by previous aerobic physical training in rats that might be, at least in part, ascribed to a decreased baroreflex sensitivity. (C) 2009 Elsevier B.V. All rights reserved.

Ultrastructural Morphology and Morphometry of Phrenic Nerve in Rats

Despite numerous literature reports on the morphometry of the myelinated fibers of phrenic nerves in rats, a systematic study of the longitudinal and lateral symmetry of the unmyelinated fibers morphometry is not available. In this study, we have undertaken ultrastructural and morphometric studies of the phrenic nerve in adult rats, assessing two different levels (proximal and distal) from both right and left sides. Phrenic nerves of adult male Wistar rats were prepared for epoxy resin embedding and transmission electron microscopy. Morphometric analysis was performed with the aid of computer software, which took into consideration the unmyelinated fiber number, density, area, and diameter, as well as ratio between myelinated and unmyelinated fibers, and the percentage of the fascicular area occupied by the myelinated and unmyelinated fibers. Comparison of data from proximal and distal segments on the same side and from the same levels between sides was performed. Differences were considered significant when P < 0.05. The most important finding is that morphometric parameters of the phrenic nerve unmyelinated fibers in adult rats are both longitudinally and laterally symmetric. This study adds important morphometric information about the unmyelinated fibers of the phrenic nerves in adult rats for proximal and distal levels on both sides of the animal. Anat Rec, 292:513-517, 2009. (C) 2008 Wiley-Liss, Inc.

Hypoalgesic effect of the transcutaneous electrical nerve stimulation following inguinal herniorrhaphy: A randomized, controlled trial

We investigated the effect of transcutaneous electrical nerve stimulation (TENS) for inguinal herniorrhaphy postoperative pain control in a prospective, randomized, double-blinded, placebo-controlled study. Forty patients undergoing unilateral inguinal herniorrhaphy with an epidural anesthetic technique were randomly allocated to receive either active TENS or placebo TENS. Postoperative pain was evaluated using a standard 10-point numeric rating scale (NRS). Analgesic requirements were also recorded. TENS (100 Hz, strong but comfortable sensory intensity) was applied for 30 minutes through 4 electrodes placed around the incision twice, 2 and 4 hours after surgery. Pain was assessed before and after each application of TENS and 8 and 24 hours after surgery. In the group treated with active TENS, pain intensity was significantly lower 2 hours (P = .028), 4 hours (P = .022), 8 hours (P = .006), and 24 hours (P = .001) after the surgery when compared with the group that received placebo TENS. Active TENS also decreased analgesic requirements in the postoperative period when compared with placebo TENS (P = .001). TENS is thus beneficial for postoperative pain relief, after inguinal herniorrhaphy; it has no observable side effects, and the pain-reducing effect continued for at least 24 hours. Consequently, the routine use of TENS after inguinal herniorrhaphy is recommended. Perspective: This study presents the hypoalgesic effect of high-frequency TENS for postoperative pain after inguinal herniorrhaphy. This may reinforce findings from basic science showing an opioid-like effect provided by TENS, given that high-frequency TENS has been shown to activate delta-opioid receptors. (C) 2008 by the American Pain Society.

Heart rate and arterial pressure variability in the experimental renovascular hypertension model in rats

This study was conducted in one kidney, one clip (1K1C) Goldblatt hypertensive rats to evaluate vascular and cardiac autonomic control using different approaches: 1) evaluation of the autonomic modulation of heart rate (HR) and systolic arterial pressure (SAP) by means of autoregressive power spectral analysis 2) assessment of the cardiac baroreflex sensitivity; and 3) double blockade with methylatropine and propranolol. The 1K1C group developed hypertension and tachycardia. The 1K1C group also presented reduction in variance as well as in LF (0.23 +/- 0.1 vs. 1.32 +/- 0.2 ms(2)) and HF (6.6 +/- 0.49 vs. 15.1 +/- 0.61 ms(2)) oscillations of pulse interval. Autoregressive spectral analysis of SAP showed that 1K1C rats had an increase in variance and LF band (13.3 +/- 2.7 vs. 7.4 +/- 1.01 mmHg(2)) in comparison with the sham group. The baroreflex gain was attenuated in the hypertensive 1K1C (- 1.83 +/- 0.05 bpm/mmHg) rats in comparison with normotensive sham (-3.23 +/- 0.06 bpm/MmHg) rats. The autonomic blockade caused an increase in the intrinsic HR and sympathetic predominance on the basal HR of 1K1C rats. Overall, these data indicate that the tachycardia observed in the 1K1C group may be attributed to intrinsic cardiac mechanisms (increased intrinsic heart rate) and to a shift in the sympathovagal balance towards cardiac sympathetic over-activity and vagal suppression associated to depressed baroreflex sensitivity. Finally, the increase in the LF components of SAP also suggests an increase in sympathetic activity to peripheral vessels. (c) 2008 Elsevier B.V. All rights reserved.

Is dystonic posturing during temporal lobe epileptic seizures the expression of an endogenous anticonvulsant system?

In temporal lobe epilepsy (TLE) seizures, tonic or clonic motor behaviors (TCB) are commonly associated with automatisms, versions, and vocalizations, and frequently occur during secondary generalization. Dystonias are a common finding and appear to be associated with automatisms and head deviation, but have never been directly linked to generalized tonic or clonic behaviors. The objective of the present study was to assess whether dystonias and TCB are coupled in the same seizure or are associated in an antagonistic and exclusive pattern. Ninety-one seizures in 55 patients with TLE due to mesial temporal sclerosis were analyzed. Only patients with postsurgical seizure outcome of Engel class I or II were included. Presence or absence of dystonia and secondary generalization was recorded. Occurrence of dystonia and occurrence of bilateral tonic or clonic behaviors were negatively correlated. Dystonia and TCB may be implicated in exclusive, non-coincidental, or even antagonistic effects or phenomena in TLE seizures. A neural network related to the expression of one behavioral response (e.g., basal ganglia activation and dystonia) might theoretically ""displace"" brain activation or disrupt the synchronism of another network implicated in pathological circuit reverberation and seizure expression. The involvement of basal ganglia in the blockade of convulsive seizures has long been observed in animal models. The question is: Do dystonia and underlying basal ganglia activation represent an attempt of the brain to block imminent secondary generalization? (C) 2007 Elsevier Inc. All rights reserved.

Microscopic Evidences That Bone Marrow Mononuclear Cell Treatment Improves Sciatic Nerve Regeneration After Neurorrhaphy

Cell therapy constitutes a possibility for improving nerve regeneration, increasing the success of nerve repair. We evaluate the use of mononuclear cells in the regeneration of the sciatic nerve after axotomy followed by end-to-end neurorrhaphy. Forty adult male Wistar rats (250300 g) were divided into four groups: (1) sham, (2) neurorrhaphy: the sciatic nerve was sectioned and repaired using epineural sutures, (3) culture medium: after the suture, received an injection of 10 mu L of culture medium into the nerve, and (4) mononuclear cell: after the suture, a concentration of 3 X 10(6) of mononuclear cell was injected in epineurium region. Mononuclear cells were obtained from the bone marrow aspirates and separated by Ficoll-Hypaque method. The histological analyses were performed at the 4th postoperative day. The sciatic functional index, histological, and morphometric analyzes were used to evaluate nerve regeneration at the 6th postoperative week. Six rats were used for immunohistochemical analysis on the 4th postoperative day. In the group 4, on the fourth day, the histological analysis demonstrated a more accelerated degenerative process and an increase of the neurotrophic factors was observed. In the 6th week, all the morphometric results of the group 4 were statistically better compared with groups 2 and 3. There was a statistically significant improvement in the sciatic functional index for group 4 compared with groups 2 and 3. Mononuclear cells stimulated nerve regeneration, most probably by speeding up the Wallerian degeneration process as well as stimulating the synthesis of neurotrophic factors. Microsc. Res. Tech. 74:355-363, 2011. (C) 2010 Wiley-Liss, Inc.

Antibothropic action of Casearia sylvestris Sw. (flacourtiaceae) extracts

Casearia sylvestris Sw., popularly known in Brazil as `guacatonga`, has been used as antitumor, antiseptic, antiulcer, local anaesthetic and healer in folk medicine. Snakebite envenomation by Bothrops jararacussu (Bjssu) constitutes a relevant public health hazard capable of inducing serious local damage in victims. This study examined the pharmacological action of apolar and polar C sylvestris leaf extracts in reverting the neuromuscular blockade and myonecrosis, which is induced by Bjssu venom and its major toxin bothropstoxin-I on the mouse phrenic nerve-diaphragm preparations. The polar methanol extract (ME) was by far the most efficacious. ME not only prevented myonecrosis and abolished the blockade, but also increased ACh release. Such facilitation in neuromuscular transmission was observed with ME alone, but was accentuated in preparations incubated with ME plus venom or toxin. This established synergy opens an interesting point of investigation because the venom or toxin in contact with ME changes from a blocking to a facilitating effect. It is suggested that rutin, known to have potent antioxidant properties, and one of the components present in the ME, could have a role in the observed effects. Since commercial rutin did not reproduce the ME effects, it is likely that a rutin-containing phytocomplex is neutralizing the bothropic envenoming effects. Copyright (C) 2008 John Wiley & Sons, Ltd.