Current status of GABA receptor subtypes in analgesia.

Gamma-aminobutyric acid (GABA), a non-protein-producing amino acid synthesized from the excitatory amino acid glutamate via the enzyme glutamic acid decarboxylase, is extensively found in microorganisms, plants and vertebrates, and is abundantly expressed in the spinal cord and brain. It is the major inhibitory neurotransmitter in the mammalian nervous system. GABA plays crucial roles in the regulation of synaptic transmission, the promotion of neuronal development and relaxation, and the prevention of insomnia and depression. As the major inhibitory neurotransmitter, GABA plays pivotal roles in the regulation of pain sensation, which is initiated by the activation of peripheral nociceptors and transmitted to the spinal cord and brain along nerves. GABA exerts these roles by directly acting on three types of receptors: ionotropic GABAA and GABAC receptors and G protein-coupled GABAB receptor. The chloride-permeable ion channel receptors GABAA and GABAC mediate fast neurotransmission, while the metabotropic GABAB receptor mediates slow effect. Different GABA receptors regulate pain sensation via different signaling pathways. Here we highlight recent updates on the involvement of specific GABA receptors and their subtypes in the process of pain sensation. Further understanding of different GABA receptors and signaling pathways in pain sensation will benefit the development of novel analgesics for pain management by targeting specific GABA receptor subtypes and signaling pathways.

Progress in the total synthesis of resin glycosides.

Resin glycosides, mainly distributed in plants of the family Convolvulaceae, are a class of novel and complex glycolipids. Their structural complexity and significant biological activities have received much attention from synthetic chemists, and a number of interesting resin glycosides have been synthesized. The synthesized resin glycosides and their analogues not only helped in structural verification, structural modification, and further biological activity exploration but also provided enlightenment for the synthesis of glycoside compounds. Herein, the present review summarizes the application of various efforts toward the synthesis of resin glycosides in the last decade.

Ras Inhibitor Lonafarnib Rescues Structural and Functional Impairments of Synapses of Aß1-42 Mice via α7nAChR-Dependent BDNF Upregulation.

Alzheimer's disease (AD) is characterized pathologically by the structural and functional impairments of synapses in the hippocampus, inducing the learning and memory deficiencies. Ras GTPase is closely related to the synaptic function and memory. This study was to investigate the effects of farnesyl transferase inhibitor lonafarnib on the synaptic structure and function in AD male mice and explore the potential mechanism. Our results showed 50 mg/kg lonafarnib (intraperitoneal) rescued the impaired spatial memory and improved the damaged synaptic transmission and plasticity of Aß1-42 mice. In addition, lonafarnib ameliorated the morphology of synaptic dendrites and spines in Aß1-42 mice. Furthermore, lonafarnib enhanced α7nAChR cell surface expression and phosphorylation of downstream Akt and CaMKII in Aß1-42 mice, which were inhibited by α7nAChR antagonist methyl lycaconitine (MLA), and increased the phosphorylation of CREB in a CaMKII- but not ERK-dependent way. Lonafarnib enhanced hippocampal brain-derived neurotrophic factor (BDNF) concentration in Aß1-42 mice, which was sensitive to MLA and KN93 (an inhibitor of CaMKII), but not related to ERK and Akt pathways. H-Ras, but not Rhes, was related to the lonafarnib induced improvement of α7nAChR cell surface expression and BDNF content. Interestingly, lonafarnib induced improvement of synaptic transmission, plasticity and spatial cognition in Aß1-42 mice was abolished by BDNF deprivation with TrkB/Fc chimera protein. Our results indicate that lonafarnib can rescue the structural and functional impairments of synapses in the Aß1-42 mice, which may be related to the improvement of BDNF content through the H-Ras-α7nAChR-dependent CaMKII-CREB pathway, leading to the improvement of spatial cognition.SIGNIFICANCE STATEMENT Alzheimer's disease (AD) is characterized pathologically by the structural and functional impairments of synapses in the hippocampus, inducing the learning and memory deficiencies. However, no effective drugs have not been developed for the treatment of AD synaptic. This study for the first time reported the beneficial effects of Ras inhibitor lonafarnib on the synaptic structure and function in AD mice, providing an alternative way for the treatment of "synaptic disease" in AD patients.

Magnolol inhibits colonic motility through down-regulation of voltage-sensitive L-type Ca2+ channels of colonic smooth muscle cells in rats.

This study aimed to investigate the effect of magnolol (5,5'-diallyl-2,2'-biphenyldiol) on contraction in distal colonic segments of rats and the underlying mechanisms. Colonic segments were mounted in organ baths for isometric force measurement. Whole-cell voltage-sensitive L-type Ca(2+) currents were recorded on isolated single colonic smooth muscle cells using patch-clamp technique. The spontaneous contractions and acetylcholine (ACh)- and Bay K 8644-induced contractions were inhibited by magnolol (3-100 µM). In the presence of Bay K8644 (100 nM), magnolol (10-100 µM) inhibited the contraction induced by 10 µM ACh. By contrast, tetrodotoxin (100 nM) and Nώ-nitro-L-arginine methyl ester (L-NAME 100 µM) did not change the inhibitory effect of magnolol (10 µM). In addition, magnolol (3-100 µM) inhibited the L-type Ca(2+) currents. The present results suggest that magnolol inhibits colonic smooth muscle contraction through downregulating L-type Ca(2+) channel activity.

Enhanced excitability and down-regulated voltage-gated potassium channels in colonic drg neurons from neonatal maternal separation rats.

UNLABELLED: Irritable bowel syndrome (IBS), characterized mainly by abdominal pain, is a functional bowel disorder. The present study aimed to examine changes in the excitability and the activity of the voltage-gated K(+) channel in dorsal root ganglia (DRG) neurons innervating the colon of rats subjected to neonatal maternal separation (NMS). Colonic DRG neurons from NMS rats as identified by FAST DiI™ labeling showed an increased cell size compared with those from nonhandled (NH) rats. Whole cell current-clamp recordings showed that colonic DRG neurons from NMS rats displayed: 1) depolarized resting membrane potential; 2) increased input resistance; 3) a dramatic reduction in rheobase; and 4) a significant increase in the number of action potentials evoked at twice rheobase. Whole cell voltage-clamp recordings revealed that neurons from both groups exhibited transient A-type (I(A)) and delayed rectifier (I(K)) K(+) currents. Compared with NH rat neurons, the averaged density of I(K) was significantly reduced in NMS rat neurons. Furthermore, the Kv1.2 expression was significantly decreased in NMS rat colonic DRG neurons. These results suggest that NMS increases the excitability of colonic DRG neurons mainly by suppressing the I(K) current, which is likely accounted for by the downregulation of the Kv1.2 expression and somal hypertrophy. PERSPECTIVE: This study demonstrates the alteration of delayed rectifier K current and Kv1.2 expression in DRG neurons from IBS model rats, representing a molecular mechanism underlying visceral pain and sensitization in IBS, suggesting the potential of Kv1.2 as a therapeutic target for the treatment of IBS.

Visceral hypersensitivity induced by activation of transient receptor potential vanilloid type 1 is mediated through the serotonin pathway in rat colon.

This study aimed to clarify the relationship between TRPV1 activation-induced visceral pain and the serotonin pathway in the colon of rats. The effects of para-chlorophenylalanine (pCPA) on visceral pain threshold pressure were assessed in capsaicin -induced visceral pain of rats. The expression of TRPV1 in the colon was examined by immunohistochemistry and Western blot analysis, and TRPV1 excitability in dorsal root ganglion (DRG) neurons was examined by whole-cell patch-clamp recording in pCPA-treated rats. Calcineurin and Ca(2+)-calmodulin-dependent kinase II (CaMKII), the important proteins in maintaining TRPV1 function in the colon, were also tested by Western blot analysis and immunofluorescence staining. Results showed that pCPA significantly increased the capsaicin-induced visceral pain threshold by 2.3-fold, and the enhanced visceral pain threshold corresponded with decreased 5-HT content (58% depleted) and enterochromaffin cell number (80% reduced). The reduced excitability of TRPV1 in DRG neurons, instead of changed TRPV1 expression, is responsible for the enhanced visceral pain threshold in 5-HT-depleted rats, and the mechanism may be related to the decreased expression of pCaMKII. These results indicate that visceral hypersensitivity induced by TRPV1 activation is modulated through 5-HT pathways and the attenuated function of TRPV1 and decreased protein expression of pCaMKII may play an important role in capsaicin-induced TRPV1 desensitization under 5-HT-depleted condition. The important role of TRPV1 and 5-HT in generating and maintaining visceral hypersensitivity may provide insights for the treatment of visceral hypersensitivity.

Inhibition of NMDA-gated ion channels by bis(7)-tacrine: whole-cell and single-channel studies.

Bis(7)-tacrine is a novel dimeric acetylcholinesterase inhibitor derived from tacrine, and has been proposed as a promising agent to treat Alzheimer's disease. We have recently reported that bis(7)-tacrine prevents glutamate-induced neuronal apoptosis by antagonizing NMDA receptors. The purpose of this study was to characterize bis(7)-tacrine inhibition of NMDA-activated current by using patch-clamp recording techniques. In cultured rat hippocampal neurons, bis(7)-tacrine inhibited NMDA-activated whole-cell current in a concentration-dependent manner with an IC(50) of 0.66+/-0.07 microM. Bis(7)-tacrine produced a gradual decline of NMDA-activated current to a steady-state, but this was not an indication of use-dependence. Also, the slow onset of inhibition by bis(7)-tacrine was not apparently due to an action at an intracellular site. Bis(7)-tacrine, 0.5 microM, decreased the maximal response to NMDA by 40% without changing its EC(50). Bis(7)-tacrine inhibition of NMDA-activated current was not voltage-dependent, and was independent of glycine concentration. Results of single-channel experiments obtained from cells expressing NR1 and NR2A subunits revealed that bis(7)-tacrine decreased the open probability and frequency of channel opening, but did not significantly alter the mean open time or introduce rapid closures. These results suggest that bis(7)-tacrine can inhibit NMDA receptor function in a manner that is slow in onset and offset and noncompetitive with respect to both NMDA and glycine. The noncompetitive inhibition of NMDA receptors by bis(7)-tacrine could contribute to its protective effect against glutamate-induced neurotoxicity.

Bis(7)-tacrine prevents glutamate-induced excitotoxicity more potently than memantine by selectively inhibiting NMDA receptors.

We have recently reported that bis(7)-tacrine could prevent glutamate-induced neuronal apoptosis through NMDA receptors. In this study, we demonstrated that in cultured rat cortical neurons, bis(7)-tacrine (IC(50), 0.02 microM) prevented glutamate-induced excitotoxicity more substantially than memantine (IC(50), 0.7 microM). In addition, bis(7)-tacrine was more efficient than memantine in buffering the intracellular Ca(2+) triggered by glutamate. In cultured rat hippocampal neurons, bis(7)-tacrine inhibited 50 microM NMDA-activated current in a concentration-dependent manner with an IC(50) of 0.68+/-0.07 microM, which is five times more potent than that produced by memantine (IC(50), 3.41+/-0.36 microM; p<0.05). By contrast, bis(7)-tacrine, up to 5 microM, did not significantly affect the current activated by 50 microM AMPA or 50 microM kainate. These results suggest that bis(7)-tacrine is more potent than memantine against glutamate-induced neurotoxicity by selectively inhibiting NMDA-activated current.

[Electrophysiological characteristic of ATP-activated currents of trigeminal ganglion neurons with different diameter in rat].

AIM: To explore the characteristic of ATP-activated current in trigeminal ganglion (TG) neurons of rat. METHODS: Whole-cell patch-clamp was performed. RESULTS: (1) The majority (92.1%) of TG neurons responded to ATP applied externally with inward currents. We recorded three distinct ATP-activated currents: fast, slow and intermediate, which were concentration-dependent. (2) In general, the fast ATP-activated currents were distributed mainly in small-diameter TG neurons, the slow ATP-activated currents were distributed mainly in large-diameter TG neurons, and the intermediate ATP-activated currents were distributed mainly in intermediate-diameter TG neurons. (3) The time course of rising phase from 10% to 90% of the three distinct ATP-activated currents were as follows: fast: (33.6 +/- 4.5) ms; intermediate: (62.2 +/- 9.9) ms; slow: (302.1 +/- 62.0) ms, and that of desensitizing phase were (399.4 +/- 58.2) ms (fast), and > 500 ms (slow) respectively. (4) From the current-voltage relationship curves, it can be seen that the reversal potential values of the three distinct ATP-activated currents were the same, all being 0-5mV. And they all were characterized by inward rectification. (5) The dose-response curve for fast ATP-activated current shifted downwards as compared with the intermediate ATP-activated current, and that for the slow ATP-activated current shifted upwards. CONCLUSION: The EC50s of the three curves tended to be identical. The results suggested that three kinds of distinct ATP-activated currents could be mediated by various subtypes of P2X receptors assembled by different subunits, and the subtypes existed in TG neurons of different diameters and transmit different information.

[Interaction of 5-HT2 and 5-HT3 receptor subtype in 5-HT-induced nociceptive responses in peripheral primary sensory nerve ending].

AIM: To study the correlation between 5-HT-induced pain response and the contribution by individual 5-HTR subtypes including 5-HT1R, 5-HT2R and 5-HT3R at the level of peripheral primary afferent. METHODS: The experiments were done on acutely isolated trigeminal ganglion (TG) neurons using whole-cell patch clamp technique and the nociceptive effect was observed on behavior experiments by intraplantar injection of test drugs. RESULTS: The majority of cells examined responded to 5-HT in a manner of concentration dependence (10(-6) - 10(-3) mol/) (61.4%, 54/88) and with a fast activating and rapid desensitizing inward current (I(5-HT)), which was thought to be mediated by the activation of 5-HT3R, since it could be blocked by 5-HT3R antagonist ICS 205930 and mimicked by 5-HT3R agonist 2-methyl-5-HT. It was found that I(5-HT) was potentiated by 5-HT2R agonist alpha-methyl-5-HT markedly, while 5-HT1R agonist R-(+)-UH 301 did not. In behavioral experiment performed on conscious rats, intraplantar injection of 5-HT(10(-5), 10(-4) and 10(-3) mol/L) induced an increment of cumulative lifting time first 20 min in a manner of concentration dependence. By dissociating 5-HTR subtypes using their corresponding antagonists (ICS and CYP) the potency order of hindpaw lifting time was identified as follows: 5-HT > 5-HT + ICS > 5-HT + CYP. CONCLUSION: The results suggest that in 5-HT-induced nociceptive response at the primary sensory level 5-HT3R may play a role of initiation, but 5-HT2R mediates maintaining and modulatory effect in the processes of nociceptive information convey.

Inhibition by bis(7)-tacrine of 5-HT-activated current in rat TG neurons.

Whole-cell recordings were performed on rat trigeminal ganglion (TG) neurons as a modeling experiment to investigate the effect of bis (7)-tacrine, a potential anti-Alzheimer's disease (AD) drug, on 5-HT-induced current (I5-HT). Extracellular 5-HT activated a concentration-dependent inward current that was blocked by ICS 205930. Co-application of bis(7)-tacrine inhibited I5-HT markedly with IC50 at 2 x 10 M. Bis(7)-tacrine shifted the concentration-response curve for I5-HT rightwards with its maximum response unchanged and EC50 increased, suggesting that this inhibition was competitive in nature. Intracellular dialysis of GDP-beta-S did not block bis(7)-tacrine inhibition of I5-HT, which excluded the involvement of G-protein mediation. These results may offer possible modality to understanding the anti-AD mechanism of bis(7)-tacrine.