Assessment of skin temperature during regional anaesthesia-What the anaesthesiologist should know.

Body temperature homeostasis is accurately regulated by complex feedback-driven neuronal mechanisms, which involve a multitude of thermoregulatory pathways. Thus, core temperature is constantly maintained within a narrow range. As one of the most effective regulatory systems skin temperature is dependent on skin blood flow. Skin blood flow in turn is highly dependent on sympathetic activity. Regional anaesthesia leads to blockade not only of somatosensory and motor nerve fibres but also of sympathetic fibres. As a consequence, vasoconstrictor tonic activity is abrogated and a vasodilation leads to an increase in skin blood flow and temperature. The aim of this review was to summarize the general physiology of thermoregulation and skin temperature as well as the alterations during regional anaesthesia. The main focus was the usefulness of measuring skin temperature as an indicator of regional anaesthesia success. According to the available literature, assessment of skin temperature can indeed serve to predict success of regional anaesthesia. Hence, it is important to realize that relevant and reliable temperature increase is only seen in the most distal body parts, ie fingers and toes. More proximally, temperature changes are frequently small and inconsistent, which means that assessment of block levels is not possible by temperature measurement. Furthermore, relevant skin temperature increases will only be observed in patients, which are initially vasoconstricted. In conclusion, measurement of skin temperature represents a reliable and feasible diagnostic tool to assess and predict the success or failure of regional anaesthesia procedures, especially in patients in which sensory testing is impossible.

Ketamine induces apoptosis via the mitochondrial pathway in human lymphocytes and neuronal cells.

BACKGROUND: Ketamine has been shown to have neurotoxic properties, when administered neuraxially. The mechanism of this local toxicity is still unknown. Therefore, we investigated the mechanism of cytotoxicity in different human cell lines in vitro. METHODS: We incubated the following cell types for 24 h with increasing concentrations of S(+)-ketamine and racemic ketamine: (i) human Jurkat T-lymphoma cells overexpressing the antiapoptotic B-cell lymphoma 2 protein, (ii) cells deficient of caspase-9, caspase-8, or Fas-associated protein with death domain and parental cells, and (iii) neuroblastoma cells (SHEP). N-Methyl-d-aspartate (NMDA) receptors and caspase-3 cleavage were identified by immunoblotting. Cell viability and apoptotic cell death were evaluated flowcytometrically by Annexin V and 7-aminoactinomycin D double staining. Mitochondrial metabolic activity and caspase-3 activation were measured. RESULTS: Ketamine, in a concentration-dependent manner, induced apoptosis in lymphocytes and neuroblastoma cell lines. Cell lines with alterations of the mitochondrial pathway of apoptosis were protected against ketamine-induced apoptosis, whereas alterations of the death receptor pathway did not reduce apoptosis. S(+)-Ketamine and racemic ketamine induced the same percentage of cell death in Jurkat cells, whereas in neuroblastoma cells, S(+)-ketamine was slightly less toxic. CONCLUSIONS: Ketamine at millimolar concentrations induces apoptosis via the mitochondrial pathway, independent of death receptor signalling. At higher concentrations necrosis is the predominant mechanism. Less toxicity of S(+)-ketamine was observed in neuroblastoma cells, but this difference was minor and therefore unlikely to be mediated via the NMDA receptor.

Apoptosis induction by different local anaesthetics in a neuroblastoma cell line.

BACKGROUND: Local anaesthetics are known to induce apoptosis in clinically relevant concentrations. Hitherto, it is unknown what determines the apoptotic potency of local anaesthetics. Therefore, we compared apoptosis induction by local anaesthetics related to their physicochemical properties in human neuronal cells. METHODS: Neuroblastoma cells (SHEP) were incubated with eight local anaesthetics, two of the ester and six of the amide types. At least, five concentrations of each local anaesthetic were evaluated. After incubation for 24 h, rates of cells in early apoptotic stages and overall cell death were evaluated by annexin V and 7-amino-actinomycin D double staining by flow cytometry. The concentrations that led to half-maximal neurotoxic effects (LD50) were calculated and compared for all local anaesthetics. RESULTS: All local anaesthetics were neurotoxic in a concentration-dependent manner. All drugs induced similar rates of early apoptotic cell formation at low concentrations, whereas at high concentrations, late apoptotic or necrotic cell death predominated. Comparison of LD50 values of the different local anaesthetics resulted in the following order of apoptotic potency from high to low toxicity: tetracaine>bupivacaine>prilocaine=mepivacaine=ropivacaine>lidocaine>procaine=articaine. The toxicity correlated with octanol/buffer coefficients and also with experimental potency of the local anaesthetic, but was unrelated to the structure (ester or amide type). CONCLUSIONS: All commonly used local anaesthetics induce neuronal apoptosis in clinically used concentrations. The neurotoxicity correlates with lipid solubility and thus with the conduction blocking potency of the local anaesthetic, but is independent of the chemical class (ester/amide).

Control of gp130 expression by the mitogen-activated protein kinase ERK2.

Interleukin (IL)-6-type cytokines such as IL-6, oncostatin M (OSM) and leukaemia inhibitory factor (LIF) signal through receptor complexes that are critically dependent on gp130. The latter is the common signal-transducing molecule that couples these cytokines to their downstream effectors, Janus kinases (JAKs) and signal transducers and activators of transcription (STATs). IL-6-type cytokine signalling additionally involves the recruitment and activation of extracellular signal-regulated kinase (ERK) 1 and ERK2. Both STATs and ERKs regulate responses mediated by members of the IL-6 family. Here, we show that ERK2, but not ERK1, also controls the expression and function of gp130 per se, as silencing ERK2 in human osteosarcoma U2OS cells inhibits the expression of gp130. This does not simply reflect quantitative differences between ERK1 and ERK2, and the effects are not restricted to osteosarcoma cells, as they can be extended to several other cancer cell types analysed to date (such as breast, prostate, lung and cervical cancer cells). Importantly, ERK2 binds to the GP130 promoter, where it perhaps interacts with the transcriptional machinery. Indeed, its role in the transcriptional regulation of the GP130 gene was corroborated using luciferase reporter assays and messenger RNA stability experiments. Considering the pivotal role that gp130 has in cancer and inflammation these data thus identify novel non-overlapping functions for ERK1 and ERK2 that are biologically relevant.

Orai1-induced store-operated Ca(2+) entry enhances phospholipase activity and modulates canonical transient receptor potential channel 6 function in murine platelets.

BACKGROUND: Orai1, the major store-operated Ca(2+) entry (SOCE) channel in platelets, is not only critical for enhancing diverse signaling pathways, but may also regulate receptor-operated Ca(2+) entry (ROCE). Dynamic coupling of the Orai1 signalosome to canonical transient receptor potential channels (TRPCs) has been suggested as an essential step in the activation of SOCE and ROCE. However, the functional significance of the biochemical interaction between Orai and TRPC isoforms remains controversial. OBJECTIVE: We aimed to elucidate the role of Orai1 in diacylglycerol (DAG)-mediated ROCE. METHODS: Trpc6(-/-) , Orai1(-/-) and Orai1(-/-) /Trpc6(-/-) mice were generated, and their platelets were analyzed. RESULTS: Thapsigargin (TG)-induced SOCE was further reduced in Orai1(-/-) /Trpc6(-/-) platelets as compared with Orai1(-/-) platelets, thus revealing that TG-induced signaling pathways can activate TRPC6. Thapsigargin-induced SOCE leads to enhanced phospholipase C and D activity in wild-type platelets. The activity of both enzymes was significantly reduced in Orai1(-/-) platelets upon TG stimulation, whereas receptor-induced phospholipase activity was not affected. Furthermore, TG-induced and glycoprotein VI-mediated thromboxane A2 release was strongly dependent on Orai1-mediated SOCE. CONCLUSION: The regulation of TRPC6 activity can occur independently of the physical interaction with Orai1. TRPC6 operates in crosstalk with Orai1 through Orai1-induced DAG production via phospholipase activation. Orai1-induced DAG production and thromboxane release amplify the second phase of Ca(2+) signaling in platelets.

Loss of spinal glycinergic neurons is not necessary for development of neuropathic pain in transgenic mice expressing enhanced green fluorescent protein in glycinergic neurons.

It has been proposed that alterations in spinal inhibitory neurotransmission are critically involved in the pathophysiology of neuropathic pain. The mechanisms by which a relief from inhibitory tone contributes to pathological pain are not fully understood. Hitherto it is still under debate whether there is a loss of inhibitory neurons in the spinal cord in neuropathic pain. The aim of the present study was to evaluate whether a specific loss of glycinergic neurons is necessary to develop hyperalgesia and allodynia in the chronic constriction injury (CCI) model of neuropathic pain. The experiments were performed in bacterial artificial chromosome (BAC) transgenic mice which specifically express enhanced green fluorescent protein under the control of the promotor of the glycine transporter 2 gene, which is a reliable marker for glycinergic neurons. Thus, possible technical inconsistencies due to immunoreactivity in fixed tissues could be ruled out. Twelve days after CCI, in neuropathic animals and in sham-operated and naive animals, lumbar and thoracic segments were analyzed using the physical disector method. Although all animals that had undergone CCI showed pathological nociceptive behavior, stereology revealed no significant difference in glycinergic neurons-neither between the different groups nor between the ipsilateral and contralateral side of the thoracic and lumbar spinal segments. Our findings suggest that a loss of glycinergic neurons is not necessary for the development of pathological nociceptive behavior in the chronic constriction injury model of neuropathic pain in mice. A different mechanism may account for the decrease in inhibitory transmission in neuropathic pain.

[Early results after varicose vein surgery--a multicenter patient inquiry]. / Frühergebnisse nach Varizenoperation--eine multizentrische Patientenbefragung als Benchmarking-Projekt der ANG.

AIM: Beside varicose vein surgery, endovenous procedures (endovenous laser therapy, VNUS and foam sclerotherapy) are now therapeutic options with the following advantages: low invasiveness, only out-patient operations, quick resumption of general activities. Whether or not classical vein surgery today still fulfills patient demands was analysed by a study group of German vascular surgeons (ANG) as a multicentre patient questioning. METHOD: The questionnaire was subdivided into eight main complexes with 24 questions about the early results of stripping operations (from start of preparation until suture removal) and the patients answers were analysed. Answers with points (1-6) and free answers were possible. RESULTS: Between 1.1.2005 and 31.3.2005 18 centres for vascular surgery collected 1708 questionnaires from patients after stripping operations. The complex "preparation and information about the operation" was scored with 1.33 to 1.39. The "general organisation" of the vascular centre was scored between 1.22 and 1.30. "Pain after operation" was rated at 1.98 (immediately), 2.26 (first day) and 2.12 (third day after operation). Reasons for other problems after surgery were haematoma (3.09), problems with the compression dressing (1.67), circulatory collapse (1.51), headache (1.33), nausea (1.25). The total score for all centres was 1.47. CONCLUSION: The early results after varicose vein surgery are good up to very good. The patients' reconvalescence time is short and there is a quick resumption of general activities. The invasiveness of vein surgery seems to be overestimated. Comparative studies to evaluate the new methods are necessary. Benchmarking projects like this study are essential parts of current quality control systems.

[Varicose vein surgery and obesity--experience with a new Boazul cuff in leg circumference up to 90 cm]. / Varizenchirurgie und Adipositas per magna--Erfahrungen mit der neuen Boazul-Manschette bis 90 cm Beinumfang.

AIM: The combination of varicose vein disease and obesity is common in clinical practice. Long operations, high blood loss and extensive bruises are the results. Since 2005, a new roll-on cuff for varicose vein surgery with a thigh circumference of 90 cm is available (big cuff). METHOD: Between 1.4.1995 and 31.3.2006, surgeons at the Clinic for Vascular Medicine, Venous and Wound Care Centre Krefeld carried out varicose vein surgery in 10 054 cases. 92% (n=9,249) of the operations were done with a roll-on cuff tourniquet. The cuff was inflated to 120 mmHg and rolled up to the thigh of the leg. We present the results of the "big cuff" pilot-study and our general experience with varicose vein surgery in bloodless technique. RESULTS: 1) Big Cuff: A transducer was located between cuff and the skin. A cuff pressure of 120 mmHg and a median thigh circumference of 79.5 cm led to a median pressure of 218.6 mmHg. The tourniquet time was 35 minutes and the systolic blood pressure during the operation did not exceed 120 mmHg. Under these conditions we did not encounter any problems. 2) Bloodless limb technique: 9 249 of our own operations using the Boazul cuff were analysed. We did not find any major complications such as acute arterial occlusion or deep vein thrombosis (DVT) during the procedure. At present the DVT rate amounts to 0.05 %. CONCLUSION: The use of the bloodless limb technique for varicose vein surgery is very comfortable, especially in cases of severe varicose veins and obesity. Maintenance of operation standards and utmost care of the cuffs lead to the successful use of this procedure.

Long-time tails of translational and rotational Brownian motion in a suspension of hard spheres.

The long-time translational and rotational Brownian motion of a sphere in a suspension of hard spheres is studied on the basis of the linearized Navier-Stokes equations and the fluctuation-dissipation theorem. It is shown that for the rotational long-time coefficient an effective medium conjecture is incorrect. There are short-range velocity correlations that decay at the same rate as the macroscopic flow pattern used in the effective medium conjecture. An estimate of the short-range correction is made on the basis of the pair term in the cluster expansion of the rotational admittance.

Cortical somatosensory-evoked potentials during spine surgery in patients with neuromuscular and idiopathic scoliosis under propofol-remifentanil anaesthesia.

BACKGROUND: Intraoperative monitoring of the spinal cord via cortical somatosensory-evoked potentials (SSEP) is a routine during spinal surgery. However, especially in neuromuscular scoliosis, the reliability of cortical SSEP has been questioned. Therefore, we compared the feasibility of cortical SSEP in idiopathic and neuromuscular scoliosis using anaesthetics known to have only minimal effect on SSEP recordings. METHODS: Total intravenous anaesthesia with propofol and remifentanil as continuous infusion was standardized for all the patients. Median and tibial nerve cortical SSEP were monitored in 54 patients who underwent surgery for spinal deformity. Twenty-seven had idiopathic scoliosis and 27 had neuromuscular scoliosis. The portion of reproducible results and intraoperative changes were compared between the groups. RESULTS: In both groups, cortical SSEP could be monitored with sufficient reliability. Only in two patients with idiopathic and four patients with neuromuscular scoliosis no reproducible traces could be obtained. The amplitudes in patients with neuromuscular scoliosis were lower than in those with idiopathic scoliosis, but not statistically significant. There were no postoperative neurological deficits. The number of false positive and true positive did not differ between the groups. CONCLUSIONS: Assessment of cortical SSEP during spine surgery was equally effective and reliable in patients with neuromuscular scoliosis and in patients with idiopathic scoliosis, possibly as a result of propofol-remifentanil anaesthesia.

Sedation during spinal anaesthesia in infants.

BACKGROUND: Neuraxial anaesthesia in adults decreases the dose of i.v. or inhalational anaesthetic needed to reach a desired level of sedation. Furthermore, spinal anaesthesia alone has a sedative effect. The mechanism behind this phenomenon is presumed to be decreased afferent stimulation of the reticular activating system after sympatholysis. We hypothesized that this mechanism is equally active in infants undergoing spinal anaesthesia. METHODS: In total, 20 unpremedicated former preterm infants underwent surgery under spinal anaesthesia with hyperbaric bupivacaine 0.5% 1 mg kg(-1) with epinephrine 10 microg kg(-1). No additional sedatives or anaesthetics were administered. Sedation was evaluated using the bispectral index (BIS) score and the 95% spectral edge frequency (SEF(95)). RESULTS: After spinal anaesthesia, mean (SD) BIS began to decrease significantly from baseline 97.0 (1.1) to 83.9 (14.4) after 15 min (P=0.006). BIS decreased further, reaching the lowest values after 30 min [62.2 (14.0); P<0.00001]. Mean (SD) SEF(95) declined from baseline 26.1 (1.8) Hz to 24.3 (3.1) after 5 min (P=0.02) and further to 9.9 (3.8) after 30 min (P<0.00001). Mean arterial pressure also decreased significantly from 66.5 (4.7) mm Hg within 10 min to 56.1 (5.6) after spinal anaesthesia (P=0.0002), while heart rate remained stable. CONCLUSIONS: These results suggest that sedation after spinal anaesthesia in infants is at least as pronounced as in adults. The sedative effect of spinal anaesthesia should be kept in mind when additional sedatives are administered, especially in former preterm infants.

Non-redundant signal transduction of interleukin-6-type cytokines. The adapter protein Shc is specifically recruited to rhe oncostatin M receptor.

The common use of the cytokine receptor gp130 has served as an explanation for the extremely redundant biological activities exerted by interleukin (IL)-6-type cytokines. Indeed, hardly any differences in signal transduction initiated by these cytokines are known. In the present study, we demonstrate that oncostatin M (OSM), but not IL-6 or leukemia inhibitory factor, induces tyrosine phosphorylation of the Shc isoforms p52 and p66 and their association with Grb2. Concomitantly, OSM turns out to be a stronger activator of ERK1/2 MAPKs. Shc is recruited to the OSM receptor (OSMR), but not to gp130. Binding involves Tyr(861) of the OSMR, located within a consensus binding sequence for the Shc PTB domain. Moreover, Tyr(861) is essential for activation of ERK1/2 and for full activation of the alpha(2)-macroglobulin promoter, but not for an exclusively STAT-responsive promoter. This study therefore provides evidence for qualitative differential signaling mechanisms exerted by IL-6-type cytokines.

A single amino acid substitution (Trp(666)-->Ala) in the interbox1/2 region of the interleukin-6 signal transducer gp130 abrogates binding of JAK1, and dominantly impairs signal transduction.

gp130 is the common signal-transducing receptor chain of interleukin (IL)-6-type cytokines. Here we describe, for the first time, a single amino acid substitution (Trp(666)-->Ala) in the membrane-proximal interbox1/2 region that abrogates activation of STAT (signal transducer and activator of transcription) transcription factors and the proliferative response of pro-B-cell transfectants. Moreover, association of the Janus kinase JAK1 is prevented. No signalling of heterodimeric IL-5 receptor (IL-5R)/gp130 chimaeras occurs in COS-7 cells, even when only a single cytoplasmic chain of a gp130 dimer contains the Trp(666)Ala mutation, indicating that it acts dominantly.

Mapping of a region within the N terminus of Jak1 involved in cytokine receptor interaction.

Janus kinase 1 (Jak1) is a cytoplasmic tyrosine kinase that noncovalently associates with a variety of cytokine receptors. Here we show that the in vitro translated N-terminal domains of Jak1 are sufficient for binding to a biotinylated peptide comprising the membrane-proximal 73 amino acids of gp130, the signal-transducing receptor chain of interleukin-6-type cytokines. By the fold recognition approach amino acid residues 36-112 of Jak1 were predicted to adopt a beta-grasp fold, and a structural model was built using ubiquitin as a template. Substitution of Tyr(107) to alanine, a residue conserved among Jaks and involved in hydrophobic core interactions of the proposed beta-grasp domain, abrogated binding of full-length Jak1 to gp130 in COS-7 transfectants. By further mutagenesis we identified the loop 4 region of the Jak1 beta-grasp domain as essential for gp130 association and gp130-mediated signal transduction. In Jak1-deficient U4C cells reconstituted with the loop 4 Jak1 mutants L80A/Y81A and Delta(Tyr(81)-Ser(84)), the interferon-gamma, interferon-alpha, and interleukin-6 responses were similarly impaired. Thus, loop 4 of the beta-grasp domain plays a role in the association of Jak1 with both class I and II cytokine receptors. Taken together the structural model and the mutagenesis data provide further insight into the interaction of Janus kinases with cytokine receptors.

A single STAT recruitment module in a chimeric cytokine receptor complex is sufficient for STAT activation.

We established a system of receptor chimeras that enabled us to induce heterodimerization of different cytoplasmic tails. Fusion constructs were created that are composed of the extracellular parts of the interleukin-5 receptor alpha and beta chains, respectively, and the transmembrane and intracellular parts of gp130, the signal transducing chain of the interleukin-6 receptor complex. In COS-7 transfectants we observed a dose-dependent interleukin-5-inducible STAT1 activation for which the presence of both the alpha and the beta chain chimera was needed. No STAT activity was detected if one of the cytoplasmic tails of the receptor complex was deleted, indicating that STAT activity resulted from a receptor dimer rather than from higher receptor aggregates. We further investigated whether dimerization of STAT1 depends on the juxtaposition of two STAT recruitment modules in a receptor complex. We show that a receptor dimer with only a single STAT1 docking site was still able to lead to STAT1 activation. This indicates that the formation of a paired set of STAT binding sites in a receptor complex is not the prerequisite for STAT factor dimerization. Our findings are discussed in view of alternative STAT dimerization models.

Contributions of leukemia inhibitory factor receptor and oncostatin M receptor to signal transduction in heterodimeric complexes with glycoprotein 130.

Leukemia inhibitory factor (LIF), cardiotrophin-1, ciliary neurotrophic factor, and oncostatin M (OSM) lead to heterodimerization of LIF receptor (LIFR) or the OSM-specific receptor (OSMR) with glycoprotein (gp) 130, the common receptor subunit for IL-6-type cytokines. Thereby intracellular signaling via Janus kinases (Jaks) and STAT transcription factors is initiated. We investigated the contributions of LIFR and OSMR to signal transduction in the context of heterodimers with gp130. Chimeric receptors based on the extracellular parts of the IL-5R alpha- and beta-chains were generated, allowing the induced heterodimerization of two different cytoplasmic tails. Our studies demonstrate that upon heterodimerization with the gp130 cytoplasmic region, the cytoplasmic parts of both LIFR and OSMR were critical for activation of an acute phase protein promoter in HepG2 hepatoma cells. The membrane-proximal region of LIFR or OSMR was crucial for the ability of such receptor complexes to induce DNA binding of STAT1 and STAT3 in COS-7 cells. Membrane-distal regions of LIFR and OSMR contributed to STAT activation even in the absence of gp130 STAT recruitment sites. We further show that the Janus kinases Jak1 and Jak2 constitutively associated with receptor constructs containing the cytoplasmic part of LIFR, OSMR, or gp130, respectively. Homodimers of the LIFR or OSMR cytoplasmic regions did not elicit responses in COS-7 cells but did in HepG2 cells and in MCF-7 breast carcinoma cells. Thus, in spite of extensive functional similarities, differential signaling abilities of gp130, LIFR, and OSMR may become evident in a cell-type-specific manner.

Signal transduction of IL-6, leukemia-inhibitory factor, and oncostatin M: structural receptor requirements for signal attenuation.

Stimulation of the IL-6R complex leads to Src homology domain containing tyrosine phosphatase 2 (SHP2) recruitment to the receptor subunit gp130 and its subsequent tyrosine phosphorylation. SHP2 is a two-SH2 domain-containing protein tyrosine phosphatase that is activated by many cytokines and growth factors. SHP2 counteracts the activation of transcription factors of the STAT family and the induction of IL-6-responsive genes. Tyrosine 759 of gp130, the signal transducing subunit of the IL-6R complex, is essential for the phosphorylation of SHP2. Mutation of tyrosine 759 to phenylalanine leads to an enhanced inducibility of IL-6-dependent genes. Here we demonstrate that no further tyrosines in the cytoplasmic part of gp130 are required for the phosphorylation of SHP2. We also tested whether the tyrosine 759 motifs in both subunits of the gp130 dimer are required for SHP2 association and tyrosine phosphorylation. Interestingly, one SHP2-recruiting phosphotyrosine motif in a single chain of the gp130 dimer is sufficient to mediate SHP2 association to the gp130 receptor subunit and its tyrosine phosphorylation as well as to attenuate IL-6-dependent gene induction. Furthermore, we show that repression of gene induction via Y759 does not require the presence of the SHP2 and STAT recruitment sites within the same receptor subunit, but within the same receptor complex. The Y759 motif in gp130 also attenuates gene induction mediated by the oncostatin M and leukemia inhibitory factor receptor complexes, which both contain gp130 as the shared subunit.