Galvanic vestibular stimulation activates the parietal and temporal cortex in humans: A functional near-infrared spectroscopy (fNIRS) study.

Galvanic vestibular stimulation (GVS) helps stabilize subjects when balance and posture are compromised. This work aimed to define the cortical regions that GVS activates in normal subjects. We used functional near-infrared spectroscopy (fNIRS) to test the hypothesis that GVS activates similar cortical areas as a passive movement. We used transcranial current stimulation (cathode in the right mastoid process and anode in the FPz frontopolar point) of bipolar direct current (2 mA), false GVS (sham), vibration (neutral stimulus), and back and forth motion (positive control of vestibular movement) in 18 clinically healthy volunteers. Seventy-two brain scans were performed, applying a crossover-type experimental design. We measured the heart rate, blood pressure, body temperature, head capacitance, and resistance before and after the experiment. The haemodynamic changes of the cerebral cortex were recorded with an arrangement of 26 channels in four regions to perform an ROI-level analysis. The back-and-forth motion produced the most significant oxygenated haemoglobin (HbO2 ) increase. The response was similar for the GVS stimulus on the anterior and posterior parietal and right temporal regions. Sham and vibrational conditions did not produce significant changes ROI-wise. The results indicate that GVS produces a cortical activation coherent with displacement percept.

Opioid receptor activation modulates the calcium current in the cochlear outer hair cells of the rat.

Previous works showed that opioid peptides are produced by olivocochlear efferent neurons, while cochlear hair cells express opioid receptors. It has been proposed that opioids protect the auditory system from damage by intense stimulation, although their use for therapeutic or illicit purposes links to hearing impairment. Therefore, it is relevant to study the effect of opioids in the auditory system to define their functional expression and mechanism of action. This study investigated the modulation of the Ca2+ currents by opioid peptides in the rat outer hair cells (OHC) using the whole-cell patch-clamp technique. The influence of agonists of the three opioid receptor subtypes (µ, δ, and κ) was studied. The κ opioid receptor agonist U-50488 inhibits the Ca2+ currents in a partially reversible form. Coincidently, norbinaltorphimine (a κ receptor antagonist) blocked the U-50488 inhibitory effect on the Ca2+ current. The δ and the µ opioid receptor agonists did not significantly affect the Ca2+ currents. These results indicate that the κ opioid receptor activation inhibits the Ca2+ current in OHC, modulating the intracellular Ca2+ concentration when OHCs depolarize. The modulation of the auditory function by opioids constitutes a relevant mechanism with a potential role in the physiopathology of auditory disturbances.

A transient decrease in heart rate with unilateral and bilateral galvanic vestibular stimulation in healthy humans.

The study of cardiovascular function with galvanic vestibular stimulation has provided evidence on the neural structures that are involved in the vestibulo-autonomic reflex. This study determined if the effect on heart rate using galvanic vestibular stimulation persists after provoking a sympathetic response and if this response differs when using unilateral or transmastoid (bilateral) stimulation. We analysed heart rate and heart rate variability using unilateral and transmastoid galvanic vestibular stimulation combined with cardiovascular reflex evoked by postural change in 24 healthy human subjects. Three electrode configurations were selected for unilateral stimulation considering the anatomical location of each semicircular canal. We compared recordings performed in seated and standing positions, and with unilateral and transmastoid stimulation. With subjects seated, a significant transient decrease in heart rate was observed with unilateral stimulation. With transmastoid stimulation, heart rate decreased in both seated and standing positions. Average intervals between normal heartbeats recorded with stimulation resemble parasympathetic cardiac function induced by auricular vagal nerve stimulation. Our results indicate that unilateral stimulation does not eliminate the natural heart rate increase caused by orthostatic hypotension. In contrast, transmastoid stimulation provoked a transient reduction in heart rate, even when subjects were standing. These responses should be considered while performing experiments with galvanic vestibular stimulation and subsequent effects in cardiac regulation mechanisms.

Nociceptin/orphanin FQ peptide receptor mediates inhibition of N-type calcium currents in vestibular afferent neurons of the rat.

The vestibular system is modulated by various neuromodulators including opioid peptides. The current study was conducted to determine whether activation of nociceptin/orphanin FQ peptide (NOP) receptors modulates voltage-gated calcium currents and action potential discharge of rat vestibular afferent neurons. We performed whole cell patch-clamp recordings on cultured vestibular afferent neurons from P7-P10 Long-Evans rats. Application of nociceptin/orphanin FQ (N/OFQ), a 17-amino acid neuropeptide that is the endogenous ligand for NOP receptor, inhibits the high-voltage activated (HVA) component of the calcium current in a concentration-dependent manner with a half inhibitory concentration of 26 nM. Said inhibitory action on the calcium current is voltage-dependent, which was made clear by the fact that it was reverted in 80% by a depolarizing prepulse. Furthermore, the effect of N/OFQ was blocked by application of the specific NOP-antagonist UFP101, by preincubation with G-protein blocker pertussis toxin, and by coapplication of the specific N-type calcium-current blocker ω-conotoxin-MVIIA. N/OFQ application causes an increase in the duration and maximum rate of repolarization of action potentials. It also decreases repetitive discharge and discharge elicited by sinusoidal stimulation. These results show that in vestibular afferents, NOP receptor activation inhibits N-type calcium current by activating G proteins, mostly through the Gßγ subunit. This suggests that NOP activation produces a presynaptic modulation of primary vestibular afferent neurons' output into the vestibular nuclei, thus taking part in the integration and gain setting of vestibular information in second-order vestibular nucleus neurons.NEW & NOTEWORTHY Our results show that in primary vestibular afferent neurons, activation of the nociceptin/orphanin FQ peptide receptor inhibits the N-type calcium current by a mechanism mediated by G proteins. We propose that calcium current inhibition modulates neurotransmitter release from vestibular afferents, producing a presynaptic modulation of vestibular input to vestibular nuclei, thus contributing to gain control in the vestibular afferent input.

Opioid modulation of cochlear auditory responses in the rat inner ear.

The auditory system has an extensive efferent innervation, which contributes to processes of control and regulation of the afferent input. The expression of receptors to various neurotransmitters and neuropeptides in the inner ear has been described, among which endogenous opioid receptors are found. The role of opioid receptors in the cochlea is not yet fully defined, it has been reported that opioid agonists and antagonists modulate the response to auditory stimuli and in clinical practice, multiple cases have been reported in which the consumption of opioid derivatives induce sensorineural hearing loss. In this work, we evaluated the effects of acute treatment with morphine, fentanyl, tramadol, and naloxone, in the auditory brain stem potentials (ABR), the compound action potential (CAP), and distortion products otacoustic emissions (DPOAE), across a wide range of stimulus frequencies and amplitudes. Adult Long-Evans rats of the strain CII/ZV weighing 180-220 g were used. For the ABR recording drugs were administered intraperitoneally or intravenously. For the CAP and DPOAE drugs were applied by direct perfusion in the middle ear. The opioid agonists produced a consistent increase in the amplitude of the PI component of the ABR and of the N1-P1 amplitude of the CAP. Naloxone produced no significant changes in the ABR and a reduction of the CAP N1-P1 amplitude. Also, opioid agonists induced a decrease in the amplitude of the DPOAE. These results show that the opioid receptor activation modulates both the afferent response at both the afferent response to acoustic stimuli, and also at the cochlear mechanics as revealed by DPOAE changes. These results present a significant step in understanding how opioid modulation of auditory responses may contribute to the auditory processing and to sensorineural hearing loss produced by opioids.

Functional diversity of secreted cestode Kunitz proteins: Inhibition of serine peptidases and blockade of cation channels.

We previously reported a multigene family of monodomain Kunitz proteins from Echinococcus granulosus (EgKU-1-EgKU-8), and provided evidence that some EgKUs are secreted by larval worms to the host interface. In addition, functional studies and homology modeling suggested that, similar to monodomain Kunitz families present in animal venoms, the E. granulosus family could include peptidase inhibitors as well as channel blockers. Using enzyme kinetics and whole-cell patch-clamp, we now demonstrate that the EgKUs are indeed functionally diverse. In fact, most of them behaved as high affinity inhibitors of either chymotrypsin (EgKU-2-EgKU-3) or trypsin (EgKU-5-EgKU-8). In contrast, the close paralogs EgKU-1 and EgKU-4 blocked voltage-dependent potassium channels (Kv); and also pH-dependent sodium channels (ASICs), while showing null (EgKU-1) or marginal (EgKU-4) peptidase inhibitory activity. We also confirmed the presence of EgKUs in secretions from other parasite stages, notably from adult worms and metacestodes. Interestingly, data from genome projects reveal that at least eight additional monodomain Kunitz proteins are encoded in the genome; that particular EgKUs are up-regulated in various stages; and that analogous Kunitz families exist in other medically important cestodes, but not in trematodes. Members of this expanded family of secreted cestode proteins thus have the potential to block, through high affinity interactions, the function of host counterparts (either peptidases or cation channels) and contribute to the establishment and persistence of infection. From a more general perspective, our results confirm that multigene families of Kunitz inhibitors from parasite secretions and animal venoms display a similar functional diversity and thus, that host-parasite co-evolution may also drive the emergence of a new function associated with the Kunitz scaffold.

Acid-Sensing Ion Channels as Potential Therapeutic Targets in Neurodegeneration and Neuroinflammation.

Acid-sensing ion channels (ASICs) are a family of proton-sensing channels that are voltage insensitive, cation selective (mostly permeable to Na+), and nonspecifically blocked by amiloride. Derived from 5 genes (ACCN1-5), 7 subunits have been identified, 1a, 1b, 2a, 2b, 3, 4, and 5, that are widely expressed in the peripheral and central nervous system as well as other tissues. Over the years, different studies have shown that activation of these channels is linked to various physiological and pathological processes, such as memory, learning, fear, anxiety, ischemia, and multiple sclerosis to name a few, so their potential as therapeutic targets is increasing. This review focuses on recent advances that have helped us to better understand the role played by ASICs in different pathologies related to neurodegenerative diseases, inflammatory processes, and pain.

Assessment of Long-Term Bowel Symptoms After Segmental Resection of Deeply Infiltrating Endometriosis: A Matched Cohort Study.

STUDY OBJECTIVE: To assess long-term bowel symptoms in women who underwent segmental bowel resection for deep-infiltrating endometriosis (DIE) compared with women who underwent resection of severe endometriosis without bowel resection. DESIGN: Cohort study with matched controls (Canadian Task Force classification II-2). SETTING: Cleveland Clinic. PATIENTS: 71 patients (36 cases and 35 controls). INTERVENTIONS: Patients who were at least 4 years out from undergoing segmental bowel resection due to DIE were matched with patients who had undergone resection of stage III/IV endometriosis without bowel resection. The patients completed validated questionnaires, and data were analyzed using the Wilcoxon rank-sum, χ(2), and Fisher exact tests. MEASUREMENTS AND MAIN RESULTS: The Bristol Stool Form Scale, Patient Assessment of Constipation Symptoms Questionnaire (PAC-SYM), and St Mark's Vaizey Fecal Incontinence Grading System were used to elicit information. The median duration of follow-up was 10.1 years (range, 4-18 years). The mean patient age and body mass index were comparable in the cases and the controls. A larger proportion of cases than controls reported new bowel symptoms (58% [21 of 36] vs 14% [5 of 35]; p = .001), as well as abdominal pain, incomplete bowel movements, and false alarms on the PAC-SYM questionnaire; however, total PAC-SYM and Vaizey Fecal Incontinence Grading System scores were similar in the 2 groups (median, 8 [interquartile range, 8-10] vs 8 [8-10]; p = .86). Similarly, the proportion of patients with normal stool consistency (Bristol Stool Form Scale score 2-6) was similar in the 2 groups (80.6% [29 of 36] vs 94.3% [33 of 35]; p = .59). CONCLUSION: Segmental bowel resection for DIE may be associated with a higher incidence of new bowel symptoms (possibly due to abdominal pain, incomplete bowel movements, and/or false alarms), but not with worse constipation or fecal incontinence, compared with surgery without bowel resection.

Supraumbilical primary trocar insertion for laparoscopic access: the relationship between points of entry and retroperitoneal vital vasculature by imaging.

OBJECTIVES: Advances in laparoscopy have demonstrated that supraumbilical primary ports can be desirable in complex cases with large masses. This study evaluated distances to vital retroperitoneal vasculature that were encountered with 45- and 90-degree angle entry from the umbilicus and 2 commonly described supraumbilical entry points at 3 and 5 cm cephalad from the umbilicus. STUDY DESIGN: Retrospective analysis of computed tomography scans of the abdomen and pelvis from 100 randomly selected women who were 18-50 years old with normal anatomy was performed. Three-dimensional models of sagittal sections were generated using IMPAX software. Measurements from the abdominal wall at the umbilicus and 3 and 5 cm cephalad with 45- and 90-degree angles to retroperitoneal structures were performed. RESULTS: With 90-degree angle entry, the abdominal wall thickness (AWT) was thinnest at the umbilicus; however, the thickness at 3 and 5 cm was similar. AWT increased at all sites with 45-degree angle entry, and the same pattern was observed. AWT and intraperitoneal distance positively correlated with body mass index and supraumbilical entry points. With 90-degree angle entry, the aorta was 1.9 cm (95% confidence interval [CI], 1.4-2.4) and 2.5 cm (95% CI, 2.0-2.9) farther away at 3 and 5 cm cephalad compared with umbilical entry. In one-third of the cases, regardless of port placement, a vascular structure other than the aorta was the most anterior vessel. With 45-degree angle entry at the umbilicus, no vessels were encountered. With 45-degree angle entry at 3 and 5 cm cephalad, the aorta was the most anterior vessel in 1% and 2% of cases, respectively, and was noted to be 1.0 cm (95% CI, 1.0-1.0) and 2.3 cm (95% CI, 1.2-3.3) farther away than with 90-degree angle entry. A vessel other than the aorta was encountered in 4% and 7% of cases at 3 and 5 cm, respectively. CONCLUSION: According to theoretic modeling, supraumbilical primary port placement can be implemented safely in laparoscopy. With supraumbilical entry, the distance to retroperitoneal vessels was greater than at the umbilicus. Compared with a 90-degree angle, with a 45-degree angle entry, it was uncommon to encounter vasculature, and all measured distances were greater.

Minimally invasive myomectomy using unidirectional knotless barbed suture.

STUDY OBJECTIVE: To demonstrate the technique for closure of the uterine wall defect during minimally invasive myomectomy using unidirectional knotless barbed suture. DESIGN: Step-by-step explanation of the technique using videos and pictures (educational video). SETTING: Women have a 70% to 80% lifetime risk of developing uterine leiomyomas. Myomectomy is a common procedure performed for conservative treatment of leiomyomas that is frequently performed using a minimally invasive technique. Knotless barbed sutures have recently been used successfully in minimally invasive myomectom procedures. Advantages of using barbed sutures in this setting include the ability to perform knotless suturing and rapid suture deployment, which may result in decreased operative time and blood loss. In addition, the tensile strength of the suture is maintained by the barbs, which facilitates the operative procedure and may potentially lead to a more even distribution of tension along the closure. INTERVENTIONS: Closure of the uterine wall defect using unidirectional knotless barbed suture during minimally invasive myomectomy. CONCLUSION: The use of unidirectional knotless barbed suture substantially facilitates closure of uterine defects during minimally invasive myomectomy and may offer additional advantages such as minimizing operative time.

Vestibular prosthesis: from basic research to clinics.

Balance disorders are highly prevalent worldwide, causing substantial disability with high personal and socioeconomic impact. The prognosis in many of these patients is poor, and rehabilitation programs provide little help in many cases. This medical problem can be addressed using microelectronics by combining the highly successful cochlear implant experience to produce a vestibular prosthesis, using the technical advances in micro gyroscopes and micro accelerometers, which are the electronic equivalents of the semicircular canals (SCC) and the otolithic organs. Reaching this technological milestone fostered the possibility of using these electronic devices to substitute the vestibular function, mainly for visual stability and posture, in case of damage to the vestibular endorgans. The development of implantable and non-implantable devices showed diverse outcomes when considering the integrity of the vestibular pathways, the device parameters (current intensity, impedance, and waveform), and the targeted physiological function (balance and gaze). In this review, we will examine the development and testing of various prototypes of the vestibular implant (VI). The insight raised by examining the state-of-the-art vestibular prosthesis will facilitate the development of new device-development strategies and discuss the feasibility of complex combinations of implantable devices for disorders that directly affect balance and motor performance.

Molecular identity, ontogeny, and cAMP modulation of the hyperpolarization-activated current in vestibular ganglion neurons.

Properties, developmental regulation, and cAMP modulation of the hyperpolarization-activated current (I(h)) were investigated by the whole cell patch-clamp technique in vestibular ganglion neurons of the rat at two postnatal stages (P7-10 and P25-28). In addition, by RT-PCR and immunohistochemistry the identity and distribution of hyperpolarization-activated and cyclic nucleotide-gated channel (HCN) isoforms that generate I(h) were investigated. I(h) current density was larger in P25-28 than P7-10 rats, increasing 410% for small cells (<30 pF) and 200% for larger cells (>30 pF). The half-maximum activation voltage (V(1/2)) of I(h) was -102 mV in P7-10 rats and in P25-28 rats shifted 7 mV toward positive voltages. At both ages, intracellular cAMP increased I(h) current density, decreased its activation time constant (τ), and resulted in a rightward shift of V(1/2) by 9 mV. Perfusion of 8-BrcAMP increased I(h) amplitude and speed up its activation kinetics. I(h) was blocked by Cs(+), zatebradine, and ZD7288. As expected, these drugs also reduced the voltage sag caused with hyperpolarizing pulses and prevented the postpulse action potential generation without changes in the resting potential. RT-PCR analysis showed that HCN1 and HCN2 subunits were predominantly amplified in vestibular ganglia and end organs and HCN3 and HCN4 to a lesser extent. Immunohistochemistry showed that the four HCN subunits were differentially expressed (HCN1 > HCN2 > HCN3 ≥ HCN4) in ganglion slices and in cultured neurons at both P7-10 and P25-28 stages. Developmental changes shifted V(1/2) of I(h) closer to the resting membrane potential, increasing its functional role. Modulation of I(h) by cAMP-mediated signaling pathway constitutes a potentially relevant control mechanism for the modulation of afferent neuron discharge.

Safety and efficacy of video laparoscopic surgical debulking of recurrent ovarian, fallopian tube, and primary peritoneal cancers.

BACKGROUND AND OBJECTIVE: Studies on the role of laparoscopy in secondary or tertiary cytoreduction for recurrent ovarian cancer are limited. Our objective is to describe our preliminary experience with laparoscopic secondary/tertiary cytoreduction in patients with recurrent ovarian, fallopian, and primary peritoneal cancers. METHODS: This is a retrospective analysis of a prospective case series. Women with recurrent ovarian, fallopian tube, or primary peritoneal cancers deemed appropriate candidates for laparoscopic debulking by the primary surgeon(s) were recruited. The patients underwent exploratory video laparoscopy, biopsy, and laparoscopic secondary/tertiary cytoreduction between June 1999 and October 2009. Variables analyzed include stage, site of disease, extent of cytoreduction, operative time, blood loss, length of hospital stay, complications, and survival time. RESULTS: Twenty-three patients were recruited. Only one surgery involved conversion to laparotomy. Seventeen (77.3%) of the patients had stage IIIC disease at the time of their initial diagnosis, and 20 (90.9%) had laparotomy for primary debulking. Median blood loss was 75 mL, median operative time 200 min, and median hospital stay 2 d. No intraoperative complications occurred. One patient (4.5%) had postoperative ileus. Eighteen (81.8%) of the patients with recurrent disease were optimally cytoreduced to 1cm. Overall, 12 patients have no evidence of disease (NED), 6 are alive with disease (AWD), and 4 have died of disease (DOD), over a median follow-up of 14 mo. Median disease-free survival was 71.9 mo. CONCLUSIONS: In a well-selected population, laparoscopy is technically feasible and can be utilized to optimally cytoreduce patients with recurrent ovarian, fallopian, or primary peritoneal cancers.

Antinociception produced by Thalassia testudinum extract BM-21 is mediated by the inhibition of acid sensing ionic channels by the phenolic compound thalassiolin B.

BACKGROUND: Acid-sensing ion channels (ASICs) have a significant role in the sensation of pain and constitute an important target for the search of new antinociceptive drugs. In this work we studied the antinociceptive properties of the BM-21 extract, obtained from the sea grass Thalassia testudinum, in chemical and thermal models of nociception in mice. The action of the BM-21 extract and the major phenolic component isolated from this extract, a sulphated flavone glycoside named thalassiolin B, was studied in the chemical nociception test and in the ASIC currents of the dorsal root ganglion (DRG) neurons obtained from Wistar rats. RESULTS: Behavioral antinociceptive experiments were made on male OF-1 mice. Single oral administration of BM-21 produced a significant inhibition of chemical nociception caused by acetic acid and formalin (specifically during its second phase), and increased the reaction time in the hot plate test. Thalassiolin B reduced the licking behavior during both the phasic and tonic phases in the formalin test. It was also found that BM-21 and thalassiolin B selectively inhibited the fast desensitizing (τ < 400 ms) ASIC currents in DRG neurons obtained from Wistar rats, with a nonsignificant action on ASIC currents with a slow desensitizing time-course. The action of thalassiolin B shows no pH or voltage dependence nor is it modified by steady-state ASIC desensitization or voltage. The high concentration of thalassiolin B in the extract may account for the antinociceptive action of BM-21. CONCLUSIONS: To our knowledge, this is the first report of an ASIC-current inhibitor derived of a marine-plant extract, and in a phenolic compound. The antinociceptive effects of BM-21 and thalassiolin B may be partially because of this action on the ASICs. That the active components of the extract are able to cross the blood-brain barrier gives them an additional advantage for future uses as tools to study pain mechanisms with a potential therapeutic application.

Alternative post-processing on a CMOS chip to fabricate a planar microelectrode array.

We present an alternative post-processing on a CMOS chip to release a planar microelectrode array (pMEA) integrated with its signal readout circuit, which can be used for monitoring the neuronal activity of vestibular ganglion neurons in newborn Wistar strain rats. This chip is fabricated through a 0.6 µm CMOS standard process and it has 12 pMEA through a 4 × 3 electrodes matrix. The alternative CMOS post-process includes the development of masks to protect the readout circuit and the power supply pads. A wet etching process eliminates the aluminum located on the surface of the p+ -type silicon. This silicon is used as transducer for recording the neuronal activity and as interface between the readout circuit and neurons. The readout circuit is composed of an amplifier and tunable bandpass filter, which is placed on a 0.015 mm2 silicon area. The tunable bandpass filter has a bandwidth of 98 kHz and a common mode rejection ratio (CMRR) of 87 dB. These characteristics of the readout circuit are appropriate for neuronal recording applications.

The aminoglycosides modulate the acid-sensing ionic channel currents in dorsal root ganglion neurons from the rat.

Acid-sensing ionic channels (ASICs) have been shown to have a significant role in a growing number of physiological and pathological processes, such as nociception, synaptic transmission and plasticity, mechanosensation, and acidosis-induced neuronal injury. The discovery of pharmacological agents targeting ASICs has significant therapeutic potential and use as a research tool. In our work, we studied the action of transient perfusion (5-15 s) of aminoglycosides (AGs) (streptomycin and neomycin) on the proton-gated ionic currents in dorsal root ganglion (DRG) neurons of the rat and in human embryonic kidney (HEK)-293 cells. In DRG neurons, streptomycin and neomycin (30 microM) produced a significant, concentration-dependent, and reversible reduction in the amplitude of the proton-gated current, and a slowing of the desensitization rate of the ASIC current. Gentamycin (30 microM) also showed a significant reversible action on the ASIC currents. The curves of the pH effect for streptomycin and neomycin indicated that their effect was not significantly affected by pH. In HEK-293 cells, streptomycin (30 microM) produced a significant reduction in the amplitude of the proton-gated current. Neomycin and gentamycin had no significant action. Reduction of extracellular Ca(2+) concentration produced a significant increase in the action of streptomycin and neomycin on the desensitization time course of ASIC currents. These results indicate that ASICs are molecular targets for AGs, which may contribute to the understanding of their actions on excitable cells. Moreover, AGs may constitute a source to develop novel molecules with a greater affinity, specificity, and selectivity for the different ASIC subunits.

Regulation of Ca v 3.1 channels by glucocorticoids.

The activity of low voltage-activated Ca(2+) (Ca(V)3) channels is tightly coupled to neurotransmitter and hormone secretion. Previous studies have shown that Ca(V)3 channels are regulated by glucocorticoids (GCs), though the mechanism underlying channel regulation remains unclear. Here, using the pituitary GH(3) cell line as a model, we investigated whether Ca(V)3 channel expression is under the control of GCs, and if their actions are mediated by transcriptional and/or post-transcriptional mechanisms. RT-PCR and western blot analyses showed that Ca(V)3.1 but not Ca(V)3.2 and Ca(V)3.3 channels is expressed in the GH(3) cells, and patch clamp recordings confirmed that Ca(2+) currents through low voltage-activated channels were decreased after chronic treatment with GCs. Consistent with this, total plasma membrane expression of Ca(V)3.1 protein as analyzed by cell-surface biotinylation assays and semi-quantitative western blotting was also down-regulated, while quantitative real-time RT-PCR analysis revealed a significant decrease of Ca(V)3.1 mRNA expression in the treated cells. In contrast, patch-clamp recordings on HEK-293 cells stably expressing recombinant Ca(V)3.1 channels showed that Ca(2+) currents were not affected by GC treatment. These results suggest that decreased transcription is a likely mechanism to explain the inhibitory actions of GCs on the functional expression of native Ca(V)3.1 channels.

Sa12b Peptide from Solitary Wasp Inhibits ASIC Currents in Rat Dorsal Root Ganglion Neurons.

In this work, we evaluate the effect of two peptides Sa12b (EDVDHVFLRF) and Sh5b (DVDHVFLRF-NH2) on Acid-Sensing Ion Channels (ASIC). These peptides were purified from the venom of solitary wasps Sphex argentatus argentatus and Isodontia harmandi, respectively. Voltage clamp recordings of ASIC currents were performed in whole cell configuration in primary culture of dorsal root ganglion (DRG) neurons from (P7-P10) CII Long-Evans rats. The peptides were applied by preincubation for 25 s (20 s in pH 7.4 solution and 5 s in pH 6.1 solution) or by co-application (5 s in pH 6.1 solution). Sa12b inhibits ASIC current with an IC50 of 81 nM, in a concentration-dependent manner when preincubation application was used. While Sh5b did not show consistent results having both excitatory and inhibitory effects on the maximum ASIC currents, its complex effect suggests that it presents a selective action on some ASIC subunits. Despite the similarity in their sequences, the action of these peptides differs significantly. Sa12b is the first discovered wasp peptide with a significant ASIC inhibitory effect.

Extracellular pH modulates the voltage-dependent Ca2+ current and low threshold K+ current in hair cells.

Extracellular protons have been shown to modulate voltage-activated ionic channels. It has been proposed that synaptic modulation by exocytosed vesicular protons would be a characteristic feature of ribbon-type synapses. Type-I hair cells have a calyceal afferent junction with a diffusionally restricted synaptic cleft. These led us to study the action of extracellular pH changes on the voltage-activated Ca(2+) and K(+) currents evaluated using a whole-cell patch clamp in isolated cells. The amplitude of the Ca(2+) and the K(+) current were reduced by extracellular acidification, but without significant changes with extracellular alkalization. A shift in the voltage dependence to a more positive membrane potential was achieved at pH < 6.8. Our results shows that the presynaptic K(+) and Ca(2+) currents are modulated by protons, indicating that protons released along with an afferent neurotransmitter would participate as a feedback mechanism in type-I hair cells.