Novel red phosphors KBaEu(XO4)3 (X = Mo, W) show high color purity and high thermostability from a disordered chained structure.

Two novel red phosphors KBaEu(XO4)3 (X = Mo, W) have been synthesized by high-temperature solid-state reactions and the crystal structures were determined for the first time. Single-crystal X-ray diffraction data reveal that their space groups are C2/c. The crystalline structure is constituted of K/BaO8 distorted square antiprisms and distorted EuO8 polyhedra which form chains lying along the c-axis and two kinds of distorted XO4 tetrahedra. This high disorder of K/Ba which might lower the crystal field symmetry around Eu3+ results in the high purity of red emission around 615 nm originating from 5D0 → 7F2 transition under near-ultraviolet (NUV) excitation. With increasing temperature, the luminescence of KBaEu(XO4)3 (X = Mo, W) phosphors decreases almost linearly with subtle alteration for the CIE coordinate. As the temperature reaches 550 K, the red emission intensity decreases to 37.3% and 50.7% of that at 300 K for KBaEu(MoO4)3 and KBaEu(WO4)3, respectively. The analysis of the decay curves of the 5D0 → 7F2 emission at variable temperatures indicates the weak cross relaxation and non-radiative energy transfer between Eu3+ ions. These results demonstrate that the investigated phosphors are attractive for application in high power NUV excited white LEDs.

Efficient diode-pumped acousto-optic Q-switched Er:Yb:GdAl(3)(BO(3))(4) pulse laser at 1522 nm.

End-pumped by a continuous-wave 976 nm diode laser, efficient 1522 nm laser operation was demonstrated in an Er:Yb:GdAl(3)(BO(3))(4) crystal when a sapphire crystal was used as a heat diffuser. A continuous-wave 1522 nm laser with a maximum output power of 750 mW and slope efficiency of 36% was realized at an absorbed pump power of 4.1 W. The pulse performances of an acousto-optic Q-switched laser with various repetition frequencies were investigated in detail. In a repetition frequency range of 1-10 kHz, 1522 nm pulse lasers with a slope efficiency of about 10%, peak output power at the kilowatt level, and width of about 50 ns were first obtained in an Er:Yb:GdAl(3)(BO(3))(4) crystal pumped by a continuous-wave diode laser. The results indicate that the crystal is a promising gain medium for an actively Q-switched 1.5 µm laser.

Nerve injury-induced calcium channel alpha-2-delta-1 protein dysregulation leads to increased pre-synaptic excitatory input into deep dorsal horn neurons and neuropathic allodynia.

BACKGROUND: Up-regulation of voltage-gated calcium channel α2 δ1 subunit post spinal nerve ligation (SNL) injury or in α2 δ1 -overexpressing transgenic (Tg) mice correlates with tactile allodynia, a pain state mediated mainly by Aß sensory fibres forming synaptic connections with deep dorsal horn (DDH) neurons. It is not clear, however, whether dysregulated α2 δ1 alters DDH synaptic neurotransmission that underlies tactile allodynia development post nerve injury. METHODS: Tactile allodynia was tested in the SNL and α2 δ1 Tg models. Miniature excitatory/inhibitory post-synaptic currents were recorded in DDH neurons from these animal models using whole-cell patch clamp slice recording techniques. RESULTS: There was a significant increase in the frequency, but not amplitude, of miniature excitatory post-synaptic currents (mEPSC) in DDH neurons that correlated with tactile allodynia in SNL and α2 δ1 Tg mice. Gabapentin, an α2 δ1 ligand that is known to block tactile allodynia in these models, also normalized mEPSC frequency dose-dependently in DDH neurons from SNL and α2 δ1 Tg mice. In contrast, neither frequency nor amplitude of miniature inhibitory post-synaptic currents was altered in DDH neurons from SNL and α2 δ1 Tg mice. CONCLUSION: Our data suggest that α2 δ1 dysregulation is highly likely contributing to tactile allodynia through a pre-synaptic mechanism involving facilitation of excitatory synaptic neurotransmission in DDH of spinal cord.

Differential effects of voltage-gated calcium channel blockers on calcium channel alpha-2-delta-1 subunit protein-mediated nociception.

BACKGROUND: Overexpression of the voltage-gated calcium channel (VGCC) alpha-2-delta1 subunit protein (Cav α2 δ1 ) has been shown to cause pain states. However, whether VGCC are involved in pain states driven by abnormal Cav α2 δ1 expression is not known. METHODS: Intrathecal injection of N-, P/Q- and L-type VGCC blockers were tested in two models: a transgenic neuronal Cav α2 δ1 overexpression (TG) model with behavioural hypersensitivity and a spinal nerve ligation (SNL) model with Cav α2 δ1 overexpression in sensory pathways and neuropathy pain states. RESULTS: The nociceptive response to mechanical stimuli was significantly attenuated in both models with ω-conotoxin GVIA (an N-type VGCC blocker) and nifedipine (an L-type VGCC blocker), in which ω-conotoxin GVIA appeared more potent than nifedipine. Treatments with ω-agatoxin IVA (P-VGCC blocker), but not ω-conotoxin MVIIC (Q-VGCC blocker) had similar potency in the TG model as the N-type VGCC blocker, while both ω-agatoxin IVA and ω-conotoxin MVIIC had minimal effects in the SNL model compared with controls. CONCLUSION: These findings suggest that, at the spinal level, N- and L-type VGCC are likely involved in behavioural hypersensitivity states driven by Cav α2 δ1 overexpression. Q-type VGCC has minimal effects in both models. The anti-nociceptive effects of P-type VGCC blocker in the Cav α2 δ1 TG mice, but minimally at the SNL model with presynaptic Cav α2 δ1 up-regulation, suggest that its potential action site(s) is at the post-synaptic and/or supraspinal level. These findings support that N-, L- and P/Q-type VGCC have differential contributions to behavioural hypersensitivity modulated by Cav α2 δ1 dysregulation at the spinal cord level.

A comprehensive, multispecialty approach to an acute exacerbation of chronic central pain in a tetraplegic.

STUDY DESIGN: We present a case report describing the multidisciplinary treatment of a tetraplegic spinal cord injury (SCI) patient who developed an acute exacerbation of chronic central pain. OBJECTIVE: To bring further awareness to the importance of using a comprehensive, multidisciplinary approach in treating acute exacerbation of chronic central pain in SCI patients. SETTING: University of California Irvine Medical Center, Orange, CA, USA. CASE REPORT: We present a 34-year-old man with a past medical history of C5 American Spinal Injury Association B tetraplegia secondary to a surfing accident 8 years prior, central pain syndrome, spasticity, autonomic dysreflexia and anxiety who arrived at the emergency room with a 1-month history of worsening acute on chronic pain refractory to opioid escalation. The multispecialty treatment plan included treatment of the patient's urinary tract infection by the primary medicine service, management of the patient's depression by the psychiatric service, treatment of bowel obstruction by general surgery and adjustment of pain medications by pain management. The patient was found to have stable neurological findings, neuroimaging unchanged from prior imaging and a urinary tract infection. Hospitalization was complicated by severe colonic dilation that required disimpaction by general surgery. CONCLUSION: The treatment of this patient's acutely worsened central pain highlights the importance of applying a multidisciplinary approach to SCI patients with an acute exacerbation of chronic central pain. In this case, the multispecialty treatment plan included treatment of the patient's urinary tract infection by the primary medicine service, management of the patient's depression by the psychiatric service, treatment of bowel obstruction by general surgery, and adjustment of pain medications by pain management.

Trigeminal nerve injury-induced thrombospondin-4 up-regulation contributes to orofacial neuropathic pain states in a rat model.

BACKGROUND: Injury to the trigeminal nerve often results in the development of chronic pain states including tactile allodynia, or hypersensitivity to light touch, in orofacial area, but its underlying mechanisms are poorly understood. Peripheral nerve injury has been shown to cause up-regulation of thrombospondin-4 (TSP4) in dorsal spinal cord that correlates with neuropathic pain development. In this study, we examined whether injury-induced TSP4 is critical in mediating orofacial pain development in a rat model of chronic constriction injury to the infraorbital nerve. METHODS: Orofacial sensitivity to mechanical stimulation was examined in a unilateral infraorbital nerve ligation rat model. The levels of TSP4 in trigeminal ganglia and associated spinal subnucleus caudalis and C1/C2 spinal cord (Vc/C2) from injured rats were examined at time points correlating with the initiation and peak orofacial hypersensitivity. TSP4 antisense and mismatch oligodeoxynucleotides were intrathecally injected into injured rats to see if antisense oligodeoxynucleotide treatment could reverse injury-induced TSP4 up-regulation and orofacial behavioural hypersensitivity. RESULTS: Our data indicated that trigeminal nerve injury induced TSP4 up-regulation in Vc/C2 at a time point correlated with orofacial tactile allodynia. In addition, intrathecal treatment with TSP4 antisense, but not mismatch, oligodeoxynucleotides blocked both injury-induced TSP4 up-regulation in Vc/C2 and behavioural hypersensitivity. CONCLUSIONS: Our data support that infraorbital nerve injury leads to TSP4 up-regulation in trigeminal spinal complex that contributes to orofacial neuropathic pain states. Blocking this pathway may provide an alternative approach in management of orofacial neuropathic pain states.

Electrophysiological characterization of spinal neuron sensitization by elevated calcium channel alpha-2-delta-1 subunit protein.

BACKGROUND: Voltage-gated calcium channel α2 δ1 subunit is the binding site for gabapentin, an effective drug in controlling neuropathic pain states including thermal hyperalgesia. Hyperalgesia to noxious thermal stimuli in both spinal nerve-ligated (SNL) and voltage-gated calcium channel α2 δ1 overexpressing transgenic (Tg) mice correlates with higher α2 δ1 levels in dorsal root ganglia and dorsal spinal cord. In this study, we investigated whether abnormal synaptic transmission is responsible for thermal hyperalgesia induced by elevated α2 δ1 expression in these models. METHODS: Behavioural sensitivities to thermal stimuli were test in L4 SNL and sham mice, as well as in α2 δ1 Tg and wild-type mice. Miniature excitatory (mEPSC) and inhibitory (mIPSC) post-synaptic currents were recorded in superficial dorsal spinal cord neurons from these models using whole-cell patch clamp slice recording techniques. RESULTS: The frequency, but not amplitude, of mEPSC in superficial dorsal horn neurons was increased in SNL and α2 δ1 Tg mice, which could be attenuated by gabapentin dose dependently. Intrathecal α2 δ1 antisense oligodeoxynucleotide treatment diminished increased mEPSC frequency and gabapentin's inhibitory effects in elevated mEPSC frequency in the SNL mice. In contrast, neither the frequency nor the amplitude of mIPSC was altered in superficial dorsal horn neurons from the SNL and α2 δ1 Tg mice. CONCLUSIONS: Our findings support a role of peripheral nerve injury-induced α2 δ1 in enhancing pre-synaptic excitatory input onto superficial dorsal spinal cord neurons that contributes to nociception development.

Thrombospondin-4 contributes to spinal cord injury-induced changes in nociception.

BACKGROUND: Our previous data have indicated that nerve injury-induced up-regulation of thrombospondin-4 (TSP4) proteins in dorsal spinal cord plays a causal role in neuropathic pain state development in a spinal nerve ligation model. To investigate whether TSP4 proteins also contribute to the development of centrally mediated changes in nociception after spinal cord injury (SCI), we investigated whether SCI induced TSP4 dysregulation, and if so, whether this change correlated with changes in nociception in a T9 spinal cord contusion injury model. METHODS: Behavioural sensitivity to mechanical, thermal stimuli and locomotor function recovery were tested blindly in SCI or sham rats post-injury. Intrathecal antisense or mismatch control oligodeoxynucleotides were used to treat SCI rats with nociceptive hyperreflexia, and Western blots were used to measure TSP4 protein levels in dorsal spinal cord samples. RESULTS: SCI induced below-level hindpaw hypersensitivity to stimuli. TSP4 protein levels are up-regulated in dorsal spinal cord of SCI rats with nociceptive hyperreflexia, but not in SCI rats without nociceptive hyperreflexia. There was no significant difference in motor function recovery post-injury between SCI rats with or without nociceptive hyperreflexia. Intrathecal treatment with TSP4 antisense, but not mismatch control, oligodeoxynucleotides led to reversal of injury-induced TSP4 up-regulation and nociceptive hyperreflexia in SCI rats. CONCLUSIONS: SCI leads to TSP4 up-regulation in lumbar spinal cord that may play a critical role in mediating centrally mediated behavioural hypersensitivity. Blocking this pathway may be helpful in management of SCI-induced changes in nociception.

Spinal 5-HT3 receptors facilitate behavioural hypersensitivity induced by elevated calcium channel alpha-2-delta-1 protein.

BACKGROUND: Peripheral nerve injury induces up-regulation of the calcium channel alpha-2-delta-1 proteins in the dorsal root ganglia and dorsal spinal cord that correlates with neuropathic pain development. Similar behavioural hypersensitivity was also observed in injury-free transgenic (TG) mice over-expressing the alpha-2-delta-1 proteins in neuronal tissues. To investigate pathways regulating alpha-2-delta-1 protein-mediated behavioural hypersensitivity, we examined whether spinal serotonergic 5-HT3 receptors are involved similarly in the modulation of behavioural hypersensitivity induced by either peripheral nerve injury in a nerve injury model or neuronal alpha-2-delta-1 over-expression in the TG model. METHODS: The effects of blocking behavioural hypersensitivity in these two models by intrathecal or systemic injections of 5-HT3 receptor antagonist, ondansetron, were compared. RESULTS: Our data indicated that the TG mice displayed similar behavioural hypersensitivities to non-painful mechanical stimulation (tactile allodynia) and painful thermal stimulation (thermal hyperalgesia) as that observed in the nerve injury model. Interestingly, tactile allodynia and thermal hyperalgesia in both models can be blocked similarly by intrathecal, but not systemic, injection of ondansetron. CONCLUSIONS: Our data suggest that spinal 5-HT3 receptors are likely to play a role in alpha-2-delta-1-mediated behavioural hypersensitivities through a descending serotonergic facilitation.

Chemotherapy-evoked painful peripheral neuropathy: analgesic effects of gabapentin and effects on expression of the alpha-2-delta type-1 calcium channel subunit.

Chemotherapeutics in the taxane and vinca-alkaloid classes sometimes produce a painful peripheral neuropathy for which there is no validated treatment. Experiments with rat models of paclitaxel- and vincristine-evoked pain suggest that these conditions may not respond to all of the analgesics that have efficacy in other models of painful peripheral neuropathy. We tested gabapentin as a potential analgesic for paclitaxel- and vincristine-evoked pain. We used a repeated dosing paradigm because there are precedents showing that repeated drug exposure may be necessary to demonstrate analgesia in neuropathic pain models. Gabapentin is believed to work via binding to voltage-gated calcium channels that contain the alpha-2-delta type-1 (alpha(2)delta-1) subunit, and the expression of this subunit is known to be increased in some painful peripheral neuropathy models. Thus we also examined whether the paclitaxel-evoked pain syndrome was accompanied by an alpha(2)delta-1 increase, and whether gabapentin had any effect on subunit expression. We found that the paclitaxel- and vincristine-evoked mechano-allodynia and mechano-hyperalgesia were significantly reduced by gabapentin, but only with repeated dosing. Paclitaxel-evoked painful peripheral neuropathy was associated with an increased expression of the alpha(2)delta-1 subunit in the spinal dorsal horn, but not in the dorsal root ganglia. The spinal cord increase was normalized by repeated gabapentin injections. Together, these findings suggest that repeated dosing with gabapentin may be beneficial in patients with chemotherapy-evoked painful peripheral neuropathy and that gabapentin's mechanisms of action may include normalization of the nerve injury-evoked increase in calcium channel alpha(2)delta-1 subunit expression.

alpha2delta and the mechanism of action of gabapentin in the treatment of pain.

Gabapentin is a drug that has been widely used in the treatment of chronic pain states. Despite its widespread usage, it is only recently that light has been shed on the mechanism of action of this agent. In the current review, the authors document the pharmacological, biochemical and molecular information that has led to the identification of the alpha2delta1 auxilliary subunit of voltage gated calcium channels as the target for this drug's actions.

Injury type-specific calcium channel alpha 2 delta-1 subunit up-regulation in rat neuropathic pain models correlates with antiallodynic effects of gabapentin.

The calcium channel alpha2delta-1 subunit is a structural subunit important for functional calcium channel assembly. In vitro studies have shown that this subunit is the binding site for gabapentin, an anticonvulsant that exerts antihyperalgesic effects by unknown mechanisms. Increased expression of this subunit in the spinal cord and dorsal root ganglia (DRG) has been suggested to play a role in enhanced nociceptive responses of spinal nerve-injured rats to innocuous mechanical stimulation (allodynia). To investigate whether a common mechanism underlies allodynic states derived from different etiologies, and if so, whether similar alpha2delta-1 subunit up-regulation correlates with these allodynic states, we compared DRG and spinal cord alpha2delta-1 subunit levels and gabapentin sensitivity in allodynic rats with mechanical nerve injuries (sciatic nerve chronic constriction injury, spinal nerve transection, or ligation), a metabolic disorder (diabetes), or chemical neuropathy (vincristine neurotoxicity). Our data indicated that even though allodynia occurred in all types of nerve injury investigated, DRG and/or spinal cord alpha2delta-1 subunit up-regulation and gabapentin sensitivity only coexisted in the mechanical and diabetic neuropathies. Thus, induction of the alpha2delta-1 subunit in the DRG and spinal cord is likely regulated by factors that are specific for individual neuropathies and may contribute to gabapentin-sensitive allodynia. However, the calcium channel alpha2delta-1 subunit is not the sole molecular change that uniformly characterizes the neuropathic pain states.

The acute antihyperalgesic action of nonsteroidal, anti-inflammatory drugs and release of spinal prostaglandin E2 is mediated by the inhibition of constitutive spinal cyclooxygenase-2 (COX-2) but not COX-1.

Western blots show the constitutive expression of COX-1 and COX-2 in the rat spinal dorsal and ventral horns and in the dorsal root ganglia. Using selective inhibitors of cyclooxygenase (COX) isozymes, we show that in rats with chronic indwelling intrathecal catheters the acute thermal hyperalgesia evoked by the spinal delivery of substance P (SP; 20 nmol) or NMDA (2 nmol) and the thermal hyperalgesia induced by the injection of carrageenan into the paw are suppressed by intrathecal and systemic COX-2 inhibitors. The intrathecal effects are dose-dependent and stereospecific. In contrast, a COX-1 inhibitor given systemically, but not spinally, reduced carrageenan-evoked thermal hyperalgesia but had no effect by any route with spinal SP hyperalgesia. Using intrathecal loop dialysis catheters, we showed that intrathecal SP would enhance the release of prostaglandin E(2) (PGE(2)). This intrathecally evoked release of spinal PGE(2) was diminished by systemic delivery of nonspecific COX and COX-2-selective inhibitors, but not a COX-1-selective inhibitor. Given at systemic doses that block SP- and carrageenan-evoked hyperalgesia, COX-2, but not COX-1, inhibitors reduced spinal SP-evoked PGE(2) release. Thus, constitutive spinal COX-2, but not COX-1, is an important contributor to the acute antihyperalgesic effects of spinal as well as systemic COX-2 inhibitors.

Upregulation of dorsal root ganglion (alpha)2(delta) calcium channel subunit and its correlation with allodynia in spinal nerve-injured rats.

Peripheral nerve injury can lead to a persistent neuropathic pain state in which innocuous tactile stimulation elicits pain behavior (tactile allodynia). Spinal administration of the anticonvulsant gabapentin suppresses allodynia by an unknown mechanism. In vitro studies indicate that gabapentin binds to the alpha(2)delta-1 (hereafter referred to as alpha(2)delta) subunit of voltage-gated calcium channels. We hypothesized that nerve injury may result in altered alpha(2)delta subunit expression in spinal cord and dorsal root ganglia (DRGs) and that this change may play a role in neuropathic pain processing. Using a rat neuropathic pain model in which gabapentin-sensitive tactile allodynia develops after tight ligation of the left fifth and sixth lumbar spinal nerves, we found a >17-fold, time-dependent increase in alpha(2)delta subunit expression in DRGs ipsilateral to the nerve injury. Marked alpha(2)delta subunit upregulation was also evident in rats with unilateral sciatic nerve crush, but not dorsal rhizotomy, indicating a peripheral origin of the expression regulation. The increased alpha(2)delta subunit expression preceded the allodynia onset and diminished in rats recovering from tactile allodynia. RNase protection experiments indicated that the DRG alpha(2)delta regulation was at the mRNA level. In contrast, calcium channel alpha(1B) and beta(3) subunit expression was not co-upregulated with the alpha(2)delta subunit after nerve injury. These data suggest that DRG alpha(2)delta regulation may play an unique role in neuroplasticity after peripheral nerve injury that may contribute to allodynia development.

Rat dorsal root ganglia express distinctive forms of the alpha2 calcium channel subunit.

Calcium channel alpha2 delta subunit is a glycosylated structural subunit consistent of the alpha2 subunit and the delta peptide. Previous studies have indicated distinctive alpha2 subunit expression in rat spinal cord and dorsal root ganglia (DRG). This study examined whether differential glycosylation underlies the molecular basis of distinct alpha2 delta subunits. The migration patterns of deglycosylated alpha2 subunits from rat spinal cord, DRG, brain and skeletal muscle were compared in Western blots. The data reported indicate that there are two forms of the alpha2 subunit in DRG that are different from the alpha2 subunit in other tissues examined, at least at the glycosylation level. Thus, post-translational modification may be important in tissue specific and functional expression of the alpha2 delta subunit.

The role of nitric oxide in nociception.

Pharmacologic, electrophysiologic, and immunohistochemical studies have suggested a role of nitric oxide (NO) in nociception processing. Recent studies have indicated that NO may modulate spinal and sensory neuron excitability through multiple mechanisms that may underlie its distinctive roles in different pain states. Differential regulation of a family of NO-producing enzymes, NO synthases, contributes mainly to the complexity underlying the role of NO in nociception. This review summarizes the latest advances in our understanding of the contribution of NO to pain transduction. Possible cellular mechanisms regarding the connection between NO production and the abnormal sensation derived from different stimuli and pathologic conditions are discussed.

Calcineurin enhances acetylcholinesterase mRNA stability during C2-C12 muscle cell differentiation.

Treatment of C2-C12 mouse myoblasts with the immunosuppressant drug cyclosporin A (CsA) enhances the increase in acetylcholinesterase (AChE) expression observed during skeletal muscle differentiation. The enhanced AChE expression is due primarily to increased mRNA stability because CsA treatment increases the half-life of AChE mRNA, but not the apparent transcriptional rate of the gene. Neither tacrolimus (FK506), an immunosuppressive agent with a distinct structure, nor cyclosporine H, an inactive congener of CsA, alters AChE expression. The enhanced AChE expression is associated with the muscle differentiation process, but cannot be triggered by CsA exposure before differentiation. Myoblasts and myotubes of C2-C12 cells express similar amounts of cyclophilin A and FKBP12, immunophilins known to be intracellular-binding targets for CsA and tacrolimus, respectively. However, cellular levels of calcineurin, a calcium/calmodulin-dependent phosphatase known to be the cellular target of ligand-immunophilin complexes, increase 3-fold during myogenesis. Overexpression of constitutively active calcineurin in differentiating cells reduces AChE mRNA levels and CsA antagonizes such an inhibition. Conversely, overexpression of a dominant negative calcineurin construct increases AChE mRNA levels, which are further enhanced by CsA. Thus, a CsA sensitive, calcineurin mediated pathway appears linked to differentiation-induced stabilization of AChE mRNA during myogenesis.

Neuronal nitric oxide synthase mRNA upregulation in rat sensory neurons after spinal nerve ligation: lack of a role in allodynia development.

Pharmacological evidence suggests a functional role for spinal nitric oxide (NO) in the modulation of thermal and/or inflammatory hyperalgesia. To assess the role of NO in nerve injury-induced tactile allodynia, we examined neuronal NO synthase (nNOS) expression in the spinal cord and dorsal root ganglia (DRG) of rats with tactile allodynia because of either tight ligation of the left fifth and sixth lumbar spinal nerves or streptozotocin-induced diabetic neuropathy. RNase protection assays indicated that nNOS mRNA (1) was upregulated in DRG, but not spinal cord, neurons on the injury side beginning 1 d after nerve ligation, (2) peaked (approximately 10-fold increase) at 2 d, and (3) remained elevated for at least 13 weeks. A corresponding increase in DRG nNOS protein was also observed and localized principally to small and occasionally medium-size sensory neurons. In rats with diabetic neuropathy, there was no significant change in DRG nNOS mRNA. However, similar increases in DRG nNOS mRNA were observed in rats that did not develop allodynia after nerve ligation and in rats fully recovered from allodynia 3 months after the nerve ligation. Systemic treatment with a specific pharmacological inhibitor of nNOS failed to prevent or reverse allodynia in nerve-injured rats. Thus, regulation of nNOS may contribute to the development of neuronal plasticity after specific types of peripheral nerve injury. However, upregulation of nNOS is not responsible for the development and/or maintenance of allodynia after nerve injury.

Splicing of 5' introns dictates alternative splice selection of acetylcholinesterase pre-mRNA and specific expression during myogenesis.

Splicing of alternative exon 6 to invariant exons 2, 3, and 4 in acetylcholinesterase (AChE) pre-mRNA results in expression of the prevailing enzyme species in the nervous system and at the neuromuscular junction of skeletal muscle. The structural determinants controlling splice selection are examined in differentiating C2-C12 muscle cells by selective intron deletion from and site-directed mutagenesis in the Ache gene. Transfection of a plasmid lacking two invariant introns (introns II and III) within the open reading frame of the Ache gene, located 5' of the alternative splice region, resulted in alternatively spliced mRNAs encoding enzyme forms not found endogenously in myotubes. Retention of either intron II or III is sufficient to control the tissue-specific pre-mRNA splicing pattern prevalent in situ. Further deletions and branch point mutations revealed that upstream splicing, but not the secondary structure of AChE pre-mRNA, is the determining factor in the splice selection. In addition, deletion of the alternative intron between the splice donor site and alternative acceptor sites resulted in aberrant upstream splicing. Thus, selective splicing of AChE pre-mRNA during myogenesis occurs in an ordered recognition sequence in which the alternative intron influences the fidelity of correct upstream splicing, which, in turn, determines the downstream splice selection of alternative exons.

The influence of Pb2+ on expression of acetylcholinesterase and the acetylcholine receptor.

This paper examines the influence of inorganic lead (Pb2+) on the presence of acetylcholinesterase (AchE) molecular forms and the acetylcholine receptor (AchR) in two types of excitable tissue, primary cultures of skeletal muscle and neural retina from embryonic chick. Treatment of skeletal muscle with Pb2+ is observed to cause reductions in the 5/7S and 19S but not the 11.4S molecular forms of AchE. The reductions are dose-dependent, requiring submicromolar concentrations, slow in onset, requiring incubation times greater than 24 hr, and tissue specific, being pronounced in skeletal muscle but absent from neural retina. Significantly, the reductions in AchE occur without corresponding reductions in amounts of AchR and without reduction in activity of protein kinase C (PKC). These studies illustrate a tissue-specific action of inorganic lead that is not mediated through PKC.