Impact of histone demethylase KDM3A-dependent AP-1 transactivity on hepatotumorigenesis induced by PI3K activation.

Epigenetic gene regulation linked to oncogenic pathways is an important focus of cancer research. KDM3A, a histone H3 lysine 9 (H3K9) demethylase, is known to have a pro-tumorigenic function. Here, we showed that KDM3A contributes to liver tumor formation through the phosphatidylinositol 3-kinase (PI3K) pathway, which is often activated in hepatocellular carcinoma. Loss of Kdm3a attenuated tumor formation in Pik3ca transgenic (Tg) mouse livers. Transcriptome analysis of pre-cancerous liver tissues revealed that the expression of activator protein 1 (AP-1) target genes was induced by PI3K activation, but blunted upon Kdm3a ablation. Particularly, the expression of Cd44, a liver cancer stem marker, was regulated by AP-1 in a Kdm3a-dependent manner. We identified Cd44-positive hepatocytes with epithelial-mesenchymal transition-related expression profiles in the Pik3ca Tg liver and confirmed their in vivo tumorigenic capacity. Notably, the number and tumor-initiating capacity of Cd44-positive hepatocytes were governed by Kdm3a. As a mechanism in Kdm3a-dependent AP-1 transcription, Kdm3a recruited c-Jun to the AP-1 binding sites of Cd44, Mmp7 and Pdgfrb without affecting c-Jun expression. Moreover, Brg1, a component of the SWI/SNF chromatin remodeling complex, interacted with c-Jun in a Kdm3a-dependent manner and was bound to the AP-1 binding site of these genes. Finally, KDM3A and c-JUN were co-expressed in 33% of human premalignant lesions with PI3K activation. Our data suggest a critical role for KDM3A in the PI3K/AP-1 oncogenic axis and propose a novel strategy for inhibition of KDM3A against liver tumor development under PI3K pathway activation.

Sharpin promotes hepatocellular carcinoma progression via transactivation of Versican expression.

Sharpin (Shank-associated RH domain-interacting protein, also known as SIPL1) is a multifunctional molecule that participates in various biological settings, including nuclear factor-κB signaling activation and tumor suppressor gene inhibition. Sharpin is upregulated in various types of cancers, including hepatocellular carcinoma (HCC), and is implicated in tumor progression. However, the exact roles of Sharpin in tumorigenesis and tumor progression remain largely unknown. Here we report novel mechanisms of HCC progression through Sharpin overexpression. In our study, Sharpin was upregulated in human HCC tissues. Increased Sharpin expression enhanced hepatoma cell invasion, whereas decrease in Sharpin expression by RNA interference inhibited invasion. Microarray analysis identified that Versican, a chondroitin sulfate proteoglycan that plays crucial roles in tumor progression and invasion, was also upregulated in Sharpin-expressing stable cells. Versican expression increased in the majority of HCC tissues and knocking down of Versican greatly attenuated hepatoma cell invasion. Sharpin expression resulted in a significant induction of Versican transcription synergistically with Wnt/ß-catenin pathway activation. Furthermore, Sharpin-overexpressing cells had high tumorigenic properties in vivo. These results demonstrate that Sharpin promotes Versican expression synergistically with the Wnt/ß-catenin pathway, potentially contributing to HCC development. A Sharpin/Versican axis could be an attractive therapeutic target for this currently untreatable cancer.

Synaptic modulation of excitatory synaptic transmission by nicotinic acetylcholine receptors in spinal ventral horn neurons.

Nicotinic acetylcholine receptors (nAChRs) are distributed widely in the central nervous system and play important roles in higher brain functions, including learning, memory, and recognition. However, functions of the cholinergic system in spinal motoneurons remain poorly understood. In this study, we investigated the actions of presynaptic and postsynaptic nAChRs in spinal ventral horn neurons by performing whole-cell patch-clamp recordings on lumbar slices from male rats. The application of nicotine or acetylcholine generated slow inward currents and increased the frequency and amplitude of spontaneous excitatory postsynaptic currents (sEPSCs). Slow inward currents by acetylcholine or nicotine were not inhibited by tetrodotoxin (TTX) or glutamate receptor antagonists. In the presence of TTX, the frequency and amplitude of miniature excitatory postsynaptic currents (mEPSCs) were also increased by acetylcholine or nicotine. A selective α4ß2 nicotinic receptor antagonist, dihydro-ß-erythroidine hydrobromide (DhßE), significantly decreased nicotine-induced inward currents without affecting the enhancement of sEPSCs and mEPSCs. In addition, a selective α7 nicotinic receptor antagonist, methyllycaconitine, did not affect either nicotine-induced inward currents or the enhancement of sEPSCs and mEPSCs. These results suggest that α4ß2 AChRs are localized at postsynaptic sites in the spinal ventral horn, non-α4ß2 and non-α7 nAChRs are located presynaptically, and nAChRs enhance excitatory synaptic transmission in the spinal ventral horn.

Reactive oxygen species enhance excitatory synaptic transmission in rat spinal dorsal horn neurons by activating TRPA1 and TRPV1 channels.

Central neuropathic pain (CNP) in the spinal cord, such as chronic pain after spinal cord injury (SCI), is an incurable ailment. However, little is known about the spinal cord mechanisms underlying CNP. Recently, reactive oxygen species (ROS) have been recognized to play an important role in CNP of the spinal cord. However, it is unclear how ROS affect synaptic transmission in the dorsal horn of the spinal cord. To clarify how ROS impact on synaptic transmission, we investigated the effects of ROS on synaptic transmission in rat spinal cord substantia gelatinosa (SG) neurons using whole-cell patch-clamp recordings. Administration of tert-butyl hydroperoxide (t-BOOH), an ROS donor, into the spinal cord markedly increased the frequency and amplitude of spontaneous excitatory postsynaptic currents (sEPSCs) in SG neurons. This t-BOOH-induced enhancement was not suppressed by the Na(+) channel blocker tetrodotoxin. However, in the presence of a non-N-methyl-D-aspartate glutamate receptor antagonist, 6-cyano-7-nitroquinoxaline-2,3-dione, t-BOOH did not generate any sEPSCs. Furthermore, in the presence of a transient receptor potential ankyrin 1 (TRPA1) channel antagonist (HC-030031) or a transient receptor potential vanilloid 1 (TRPV1) channel antagonist (capsazepine or AMG9810), the t-BOOH-induced increase in the frequency of sEPSCs was inhibited. These results indicate that ROS enhance the spontaneous release of glutamate from presynaptic terminals onto SG neurons through TRPA1 and TRPV1 channel activation. Excessive activation of these ion channels by ROS may induce central sensitization in the spinal cord and result in chronic pain such as that following SCI.

Pilocytic astrocytomas in elderly adults.

Pilocytic astrocytomas are classified as WHO grade I gliomas that occur predominantly in children and young adults. Reports of the tumors in elderly adults are extremely rare. We describe two cases of pilocytic astrocytoma in elderly adults, a 68-year-old man and a 71-year-old woman. Brain computed tomography (CT) and magnetic resonance imaging (MRI) revealed well-circumscribed lesions associated with contrast enhancement and minimal surrounding edema. Pathological studies revealed findings consistent with pilocytic astrocytomas. Although these tumors are rarely found in elderly adults, pilocytic astrocytomas should be considered in the differential diagnosis if the radiographic features of the tumors are characteristic of pilocytic astrocytomas.

Effect of resiniferatoxin on glutamatergic spontaneous excitatory synaptic transmission in substantia gelatinosa neurons of the adult rat spinal cord.

The transient receptor potential (TRP) vanilloid type 1 (TRPV1) agonist, capsaicin, enhances glutamatergic spontaneous excitatory synaptic transmission in CNS neurons. Resiniferatoxin (RTX) has a much higher affinity for TRPV1 than capsaicin, but its ability to modulate excitatory transmission is unclear. We examined the effect of RTX on excitatory transmission using the whole-cell patch-clamp technique in substantia gelatinosa (SG) neurons of adult rat spinal cord slices. Bath-applied RTX dose-dependently increased the frequency, but not the amplitude, of spontaneous excitatory postsynaptic current (sEPSC), independent of its application time. In about a half of the neurons tested, this effect was accompanied by an inward current at -70 mV that was sensitive to glutamate-receptor antagonists. Repeated application of RTX did not affect excitatory transmission. RTX was more potent than capsaicin but showed similar efficacy. RTX activity could be blocked by capsazepine or SB-366791, a TRPV1 antagonist, but not tetrodotoxin, a Na(+)-channel blocker, and could be inhibited by pretreatment with capsaicin but not the TRPA1 agonist, allyl isothiocyanate. RTX enhances the spontaneous release of L-glutamate from nerve terminals with similar efficacy as capsaicin and produces a membrane depolarization by activating TRPV1 in the SG, with fast desensitization and slow recovery from desensitization. These results indicate a mechanism by which RTX can modulate excitatory transmission in SG neurons to regulate nociceptive transmission.

Proteinase-activated receptor-1 activation presynaptically enhances spontaneous glutamatergic excitatory transmission in adult rat substantia gelatinosa neurons.

Proteinase-activated receptors (PARs) have a unique activation mechanism in that a proteolytically exposed N-terminal region acts as a tethered ligand. A potential impact of PAR on sensory processing has not been fully examined yet. Here we report that synthetic peptides with sequences corresponding to PAR ligands enhance glutamatergic excitatory transmission in substantia gelatinosa (SG) neurons of adult rat spinal cord slices by using the whole cell patch-clamp technique. The frequency of spontaneous excitatory postsynaptic current (EPSC) was increased by PAR-1 agonist SFLLRN-NH2 (by 47% at 1 microM) with small increases by PAR-2 and -4 agonists (SLIGKV-NH2 and GYPGQV-OH, respectively; at >3 microM); there was no change in its amplitude or in holding current at -70 mV. The PAR-1 peptide action was inhibited by PAR-1 antagonist YFLLRNP-OH. TFLLR-NH2, an agonist which is more selective to PAR-1 than SFLLRN-NH2, dose-dependently increased spontaneous EPSC frequency (EC50=0.32 microM). A similar presynaptic effect was produced by PAR-1 activating proteinase thrombin in a manner sensitive to YFLLRNP-OH. The PAR-1 peptide action was resistant to tetrodotoxin and inhibited in Ca2+-free solution. Primary-afferent monosynaptically evoked EPSC amplitudes were unaffected by PAR-1 agonist. These results indicate that PAR-1 activation increases the spontaneous release of L-glutamate onto SG neurons from nerve terminals in a manner dependent on extracellular Ca2+. Considering that sensory processing within the SG plays a pivotal role in regulating nociceptive transmission to the spinal dorsal horn, the PAR-1-mediated glutamatergic transmission enhancement could be involved in a positive modulation of nociceptive transmission.

Elastic plasma protein film blended with platelet releasate accelerates healing of diabetic mouse skin wounds.

BACKGROUND AND OBJECTIVES: The growth factors derived from platelets and plasma proteins mediate the wound-healing process that is characterized by the sequential migration and differentiation of several cell populations that give rise to angiogenesis, collagen synthesis, wound contraction, and re-epithelialization. To evaluate the efficacy of the blood-derived factors in wound healing, we examined a novel wound dressing consisting of concentrated human plasma proteins and platelet releasate (CPPP). MATERIALS AND METHODS: To generate CPPP, plasma proteins and platelets in the peripheral blood (n = 5) were concentrated with the cold ethanol precipitation method. The thrombin obtained from the same blood unit and calcium chloride (CaCl(2)) were mixed to a concentrate. The CPPP has enough strength to dress cutaneous wounds and contains large amounts of cytokines and fibronectin. We applied the CPPP to excisional skin wounds in genetically healing-impaired model mice (n= 5) and the wounds were evaluated 10 days after the operation. RESULTS: The area of CPPP-treated wounds decreased significantly compared with that of the control wounds (65% vs. 94% of the original size, respectively, P= 0.032). The immunostained section revealed a striking effect of CPPP on vascularization compared with the control wounds (13.2 vs. 2.7 vessels per mm(2) as mean vascular density observed in the sections, respectively, P= 0.013). CONCLUSIONS: Our results suggest that CPPP is a promising biologically active dressing for full-thickness skin wounds. CPPP can be an entirely autologous biological dressing, suggesting that it is free from the risk of transmission of pathogens through blood products.

Tramadol, but not its major metabolite (mono-O-demethyl tramadol) depresses compound action potentials in frog sciatic nerves.

BACKGROUND AND PURPOSE: Although tramadol is known to exhibit a local anaesthetic effect, how tramadol exerts this effect is not understood fully. EXPERIMENTAL APPROACH: The effects of tramadol and its metabolite mono-O-demethyl-tramadol (M1) on compound action potentials (CAPs) were examined by applying the air-gap method to frog sciatic nerves, and the results were compared with those of other local anaesthetics, lidocaine and ropivacaine. KEY RESULTS: Tramadol reduced the peak amplitude of the CAP in a dose-dependent manner (IC50=2.3 mM). On the other hand, M1 (1-2 mM), which exhibits a higher affinity for mu-opioid receptors than tramadol, did not affect CAPs. These effects of tramadol were resistant to the non-selective opioid receptor antagonist naloxone and the mu-opioid receptor agonist, DAMGO, did not affect CAPs. This tramadol action was not affected by a combination of the noradrenaline uptake inhibitor, desipramine, and the 5-hydroxytryptamine uptake inhibitor, fluoxetine. Lidocaine and ropivacaine also concentration-dependently reduced CAP peak amplitudes with IC50 values of 0.74 and 0.34 mM, respectively. CONCLUSIONS AND IMPLICATIONS: These results indicate that tramadol reduces the peak amplitude of CAP in peripheral nerve fibres with a potency which is less than those of lidocaine and ropivacaine, whereas M1 has much less effect on CAPs. This action of tramadol was not produced by activation of mu-opioid receptors nor by inhibition of noradrenaline and 5-hydroxytryptamine uptake. It is suggested that the methyl group present in tramadol but not in M1 may play an important role in producing nerve conduction block.

Evaluating dressing materials for the prevention of shear force in the treatment of pressure ulcers.

OBJECTIVE: Shear force is believed to affect pressure ulceration. Therefore, dressing materials that reduce shear force may prevent ulceration and facilitate healing. METHOD: We measured the following three properties: the coefficient of friction between the outer layer of the dressings and the patient's clothes; the degree of adhesiveness between the inner layer of the dressing and the patient's skin; the transmissibility of shear force of the dressing. RESULTS: The coefficients of static friction were 1.01 for hydropolymer, 0.72 for hydrofoam and 0.48 for hydrocolloid. Adhesiveness was tested by rolling different sized ball bearings down a slope and over the adhesive lining under both wet and dry conditions. Under dry conditions, the heaviest ball bearing that stopped rolling for five seconds was 111.9g for both hydrofoam and hydrocolloid. Under wet condition, it was 11.9g for hydrofoam and under 1g for hydrocolloid. Tests showed the very low transmissibility (I N buffer) of shear force for hydrofoam, with significant differences between the dressings. Clinical observation has identified good results for hydrofoam when used under highly exuding conditions and for hydrocolloid when used with relatively slight or decreased exudate. CONCLUSION: Existing dressing materials are being developed and evaluated for wound healing. However, if innovations in the raw materials from which dressings are manufactured could lead to a reduction in shear force and the prevention of pressure ulcers, then dressing materials could be discussed from a viewpoint that is quite different from wound healing.

Benefits of surgical reconstruction in pressure ulcers with a non-advancing edge and scar formation.

OBJECTIVE: To highlight the difficulty in treating pressure ulcers associated with scar formation and the benefits of surgical treatment in these cases by presenting a series of clinical cases. METHOD: Nine sacral ulcers surrounded by significant scar formation and 14 sacral ulcers without significant scars were treated using the same conservative management protocol. Healing status was quantitatively estimated using an assessment tool called DESIGN, which classifies pressure ulcer severity and allows healing to be monitored; a decreasing score indicates progressive improvement. RESULTS: Ulcers with scar formation showed little response to the treatment, while those without remarkable scars exhibited a significant decrease of the DESIGN scores. Surgical reconstruction was conducted in five ulcers surrounded by scar formation, and all showed complete healing within extremely short periods. CONCLUSION: Conservative treatment of pressure ulcers with a non-advancing edge surrounded by scar formation presents considerable difficulties. Surgical reconstruction offers many benefits for non-healing ulcers.

The positive experience of using a growth factor product on deep wounds with exposed bone.

Trafermin, a form of basic fibroblast growth factor, has been used in Japan since 2001. This study investigates whether it can facilitate closure in wounds with deep soft-tissue defects and exposed bone, where surgical closure is not possible.

Sprouting from arteriovenous shunt vessels with increased blood flow.

The purpose of this study was to evaluate the effect of increased blood flow on angiogenesis at the large vessels. The arteriovenous (AV) shunt was made on the thigh of male Wistar rats (n = 27) to increase blood flow, wrapped with artificial skin dermis, which consisted of a silicon outer layer, and isolated from surrounding tissues. Blood flow increased from 2.40 +/- 0.77 to 35.8 +/- 8.7 ml min(-1) (14.9 times), and the shear stress index (relative value of shear stress) increased from 10.7 +/- 3.6 to 73.4 +/- 18.1 (6.85 times) 60 min after the shunt formation. Newly formed vessels were observed around the AV shunt loop. Scanning electron micrographs at the AV shunt vessel lumen showed modified endothelial cells at day 7 and a remarkable number of pores at day 14. The volume of newly formed vessels was increased 12 times from day 5 to day 14. The mechanical factor of shear stress was considered the major stimulator of angiogenesis. This is the first report of electron-microscopic observation of sprouts from a large vessel lumen. The new AV shunt model is useful for basic research on angiogenesis at the large vessels in vivo and, furthermore, could generate vascularised tissues with various cultured cells.

Functional reorganization of the spinal pain pathways in developmental and pathological conditions.

Following inflammation, a subpopulation of Abeta afferents that terminates preferentially in deeper laminae have been shown to extend their axons to the superficial dorsal horn, particularly substantia gelatinosa (SG). Similarly, SG neurons in immature spinal cord receive mainly Abeta afferent inputs. To clarify whether the reorganized sensory pathway in the inflamed rats has a functional similarity with that in the developmental state, we compared synaptic inputs from primary afferents using in vitro and in vivo patch-damp recordings from SG neurons. SG neurons in the mature state had monosynaptic inputs from Adelta and C afferents, while only a few neurons received inputs from Abeta afferents. Following inflammation, the Abeta afferents extended their axons to SG and established functional monosynaptic transmission. Meanwhile, SG neurons in the immature state received preferentially Abeta as well as Adelta afferent inputs, and the majority of Abeta afferent inputs were monosynaptic. These observations support the idea that the sprouting of the large afferent fibres observed in inflamed rats is, at least in part, a regeneration process. However, the process, maybe distinct at some point from the process during development, therefore, produces pathological pain. Though the idea that the regeneration mimics the developmental process has been widely accepted, other possibilities cannot be excluded.

Analysis of receptive fields revealed by in vivo patch-clamp recordings from dorsal horn neurons and in situ intracellular recordings from dorsal root ganglion neurons.

It has been thought that spinal dorsal horn neurons receive convergent inputs from not only somatosensory but also visceral pathways. For instance, the referred pain is presumed to be due to the convergence of sensory inputs from cardiac and shoulder receptive fields. However, precise investigation has not been made from dorsal horn neurons yet, because of difficulty in studying the pathways from those regions by means of conventional electrophysiology. The purpose of this study is to clarify the convergent inputs to single dorsal horn neurons from wide receptive fields using an in vivo patch-clamp recording technique from the superficial spinal dorsal horn and an intracellular recording from dorsal root ganglion neurons that keep physiological connections with the peripheral sites. Identified dorsal root ganglion neurons received an input from a quite small area, about 1 x 1 mm in width of the skin. In contrast, substantia gelatinosa neurons in the spinal cord received inputs from an unexpectedly wide area of the skin. Previous extracellular recordings have, however, revealed that substantia gelatinosa neurons have small receptive field. This discrepancy is probably due mainly to an availability of the in vivo patch-clamp method to analyze sub-threshold synaptic responses. In contrast, the extracellular recording technique allows us to analyze predominantly the firing frequency of neurons. Thus, the in vivo patch-clamp recordings from dorsal horn neurons and the intracellular recordings from DRG neurons will be useful for well understanding the sensory processing in the spinal cord.

Depletion of substance P from rat primary sensory neurons by ATP, an implication of P2X receptor-mediated release of substance P.

Effects of ATP on substance P immunoreactivity were examined in cultured dorsal root ganglion neurons. We found that treatment of dorsal root ganglion neurons with ATP significantly depleted substance P immunoreactivity on the neurites and somata of the neurons. The effects of ATP were significantly inhibited by the purinergic P2 receptor antagonists suramin (30 microM) and pyridoxal-phosphate-6-azophenyl-2',4'-disulfonic acid (10 microM). We also showed that ATP-induced depletion of substance P immunoreactivity from dorsal root ganglion neurons depended on the entry of Ca(2+). In a spinal cord slice preparation, we also found the internalization of neurokinin-1/substance P receptors in many dorsal horn neurons following the application of ATP or alpha,beta-methylene-ATP. Together these results indicate that activation of P2X receptors may result in release of substance P from primary afferent neurons.

Clinical use of amrinone (a selective phosphodiesterase III inhibitor) in reconstructive surgery.

Amrinone is a selective phosphodiesterase III inhibitor that increases cyclic adenosine monophosphate by preventing its breakdown. It is effective in the treatment of congestive heart failure because of its ability to increase myocardial contractility and vascular smooth muscle relaxation. This study was designed to clarify the potential efficacy of amrinone in plastic surgery by clinically assessing its ability to enhance flap blood flow after reconstructive surgery and relieve intraoperative vasospasm. Its effects were compared with those of prostaglandin E1 and lidocaine, which are widely approved agents for improving the hemodynamics of flaps. In the first clinical study, the effects on flap blood flow after flap transfers were investigated. Twenty-six patients underwent reconstructive surgery with vascularized free or pedicled flaps. Blood flow was measured before and 60 minutes after intravenous infusion of lactated Ringer solution (control), amrinone (10 microg/kg/min), or prostaglandin E1 (10 ng/kg/min) using a laser Doppler flowmeter. In the second study, the effects on relief of vasospasm during operation were evaluated. The blood flow of 28 island flaps was measured by laser Doppler flowmetry immediately after flap elevation and 10 minutes after topical application of saline (control), amrinone (5 mg/ml), or lidocaine (10%) to the pedicle in an attempt to resolve the vasospasm. In both clinical studies, the effects of amrinone were statistically no less than those of prostaglandin E1 and lidocaine. The results show that amrinone positively influences the microcirculatory blood flow of transferred flaps and relieves intraoperative vasospasm in clinical cases. The present study suggests that amrinone could be useful for postoperative and intraoperative care in reconstructive surgery.

Mandibular reconstruction using microvascular free flaps: a statistical analysis of 178 cases.

For this article, 178 consecutive cases of mandibular reconstruction using microvascular free flaps and performed from 1979 to 1997 were studied. The purpose of this report is to compare flap success rates, complications, and aesthetic and functional results. The ages of the 131 men and 47 women ranged from 13 to 85 years, with an average of 55 years. Donor sites included the rib (11 cases), radius (one case), ilium (36 cases), scapula (51 cases), fibula (34 cases), and soft-tissue flaps with implant (45 cases). Complications included total flap necrosis, partial flap necrosis, major fistula formation, and minor fistula formation. The rate of total flap necrosis involving the ilium and fibula was significantly higher than that of all other materials combined (p < 0.05). The overall rate of implant plate removal, which resulted from the exposure or fracture of the plate, was 35.6 percent (16 of 45 cases). Each mandibular defect was classified by the extent of the bony defect and by the extent of the soft-tissue defect. The extent of the mandibular bony defect was classified according to the HCL method of Jewer et al. The extent of the soft-tissue defect was classified into four groups: none, skin, mucosal, and through-and-through. According to these classifications, functional and aesthetic assessments of deglutition and contour were performed on 115 subjects, and speech was evaluated in 110. To evaluate the postoperative results, points were assigned to each assessment of deglutition, speech, and mandibular contour. Statistical analysis between pairs of bone-defect groups revealed that there was no significant difference in each category. Regarding deglutition, statistical analysis between pairs of soft-tissue-defect groups revealed there were significant differences (p < 0.05) between the none and the mucosal groups and also between the none and the through-and-through groups. Regarding speech, there was a significant difference (p < 0.05) between the none and the through-and-through groups. Regarding contour, there were significant differences (p < 0.01) between the none and the through-and-through groups and between the mucosal and the through-and-through groups. The points given for each function, depending on the reconstruction material, revealed that there was no significant difference between pairs of material groups. From this prospective study, the authors have developed an algorithm for oromandibular reconstruction. When the bony defect is lateral, the ilium, fibula, or scapula should be chosen as the donor site, depending on the extent of the soft-tissue defect. When the bony defect is anterior, the fibula is always the best choice. When the soft-tissue defect is extensive or through-and-through with an anterior bony defect, the fibula should be used with other soft-tissue flaps.

Deep circumflex iliac perforator flap with iliac crest for mandibular reconstruction.

The deep circumflex iliac myocutaneous perforator (DCIP) flap with iliac crest was used to reconstruct oromandibular defects in 10 patients. In seven of the patients a dominant perforator was found preoperatively using a Doppler flowmeter; in five of these seven patients a DCIP flap was successfully transferred. In two of the seven patients the dominant perforators were too narrow: one patient underwent a standard osteocutaneous flap transfer and one patient underwent a second flap transfer. In three patients no dominant perforator was found before or during surgery. The freedom of the DCIP flap from the harvested iliac crest facilitates correct positioning. However, to ensure that the DCIP flap can be safely elevated, the presence of perforators must be confirmed preoperatively. Even when a perforator has been identified, complicated dissection may be necessary. We stress the importance of a thorough knowledge of the anatomy of second flaps and of obtaining informed consent to use them.

ATP P2X receptor-mediated enhancement of glutamate release and evoked EPSCs in dorsal horn neurons of the rat spinal cord.

Presynaptic ATP P2X receptors have been proposed to play a role in modulating glutamate release from the first sensory synapse in the spinal cord. Using spinal cord slice preparations and patch-clamp recordings from dorsal horn neurons in lamina V of the rat spinal cord, we showed that the activation of P2X receptors by alpha,beta-methylene-ATP (alphabetam-ATP) resulted in a large increase in the frequency of spontaneous EPSCs (sEPSCs) and miniature EPSCs (mEPSCs). The increases in mEPSC frequency by alphabetam-ATP were not blocked by the Ca(2+) channel blocker, 30 microm La(3+), but were abolished in a bath solution when Ca(2+) was omitted. The increases in mEPSC frequency by alphabetam-ATP were blocked completely by the P2 receptor antagonist pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS) at 10 microm. Furthermore, the EPSCs evoked by dorsal root stimulation were potentiated by alphabetam-ATP as well as by the ecto-ATPase inhibitor ARL67156 and were depressed in the presence of P2 receptor antagonists PPADS (10 microm) and suramin (5 microm). The effects of these compounds on the evoked EPSCs were associated with the changes in glutamate release probability of primary afferent central terminals. Our results indicate that alphabetam-ATP-sensitive P2X receptors play a significant role in modulating excitatory sensory synaptic transmission in the spinal cord, and the potential role of endogenous ATP is suggested.