Cryopreservation of autologous bone flaps following decompressive craniectomy: A new method reduced positive cultures without increase in post-cranioplasty infection rate.

Introduction: Cranioplasty (CP) after decompressive craniectomy (DC) is a common neurosurgical procedure. Implementation of European Union (EU) directives recommending bacterial cultures before cryopreservation, lead to increased number of autologous bone flaps being discarded due to positive cultures. A new method for handling bone flaps prior to cryopreservation, including the use of pulsed lavage, was developed. Research question: The aim was to evaluate the effect of a new method on proportion of positive bacterial cultures and surgical site infection (SSI) following CP surgery. Material and methods: Sixty-one bone flaps from 53 consecutive DC surgery patients were retrospectively included and the study period was divided into before and after method implementation. Patient demographics, laboratory and culture results, type of CP and occurrence of SSI were analyzed. Results: Twenty-six and 18 bone flaps were available for analysis during the first and second period, respectively. The proportion of positive bacterial cultures was higher in the first period compared to the second (n â€‹= â€‹9(35%) vs 0(0%); p â€‹= â€‹0.001), and thus the use of custom made implants was considerably higher in the first study period (p â€‹= â€‹0.001). There was no difference in the frequency of post-cranioplasty SSI between the first and second study period (n â€‹= â€‹3 (11.5%) vs 1 (4.8%), p â€‹= â€‹0.408). Discussion and conclusion: The new method for handling bone flaps resulted in a lower frequency of positive bacterial cultures, without increased frequency of post-cranioplasty SSI, thus demonstrating it is safe to use, allows compliance with the EU-directives, and may reduce unnecessary discarding of bone flaps.

Connexin36 knockout mice display increased sensitivity to pentylenetetrazol-induced seizure-like behaviors.

OBJECTIVE: Large-scale synchronous firing of neurons during seizures is modulated by electrotonic coupling between neurons via gap junctions. To explore roles for connexin36 (Cx36) gap junctions in seizures, we examined the seizure threshold of connexin36 knockout (Cx36KO) mice using a pentylenetetrazol (PTZ) model. METHODS: Mice (2-3months old) with Cx36 wildtype (WT) or Cx36KO genotype were treated with vehicle or 10-40mg/kg of the convulsant PTZ by intraperitoneal injection. Seizure and seizure-like behaviors were scored by examination of video collected for 20min. Quantitative real-time PCR (QPCR) was performed to measure potential compensatory neuronal connexin (Cx30.2, Cx37, Cx43 and Cx45), pannexin (PANX1 and PANX2) and gamma-aminobutyric acid type A (GABA(A)) receptor α1 subunit gene expression. RESULTS: Cx36KO animals exhibited considerably more severe seizures; 40mg/kg of PTZ caused severe generalized (≥grade III) seizures in 78% of KO, but just 5% of WT mice. A lower dose of PTZ (20mg/kg) induced grade II seizure-like behaviors in 40% KO vs. 0% of WT animals. There was no significant difference in either connexin, pannexin or GABA(A) α1 gene expression between WT and KO animals. CONCLUSION: Increased sensitivity of Cx36KO animals to PTZ-induced seizure suggests that Cx36 gap junctional communication functions as a physiological anti-convulsant mechanism, and identifies the Cx36 gap junction as a potential therapeutic target in epilepsy.

GABAergic compensation in connexin36 knock-out mice evident during low-magnesium seizure-like event activity.

Gap junctions within the cerebral cortex may facilitate cortical seizure formation by their ability to synchronize electrical activity. To investigate this, one option is to compare wild-type (WT) animals with those lacking the gene for connexin36 (Cx36 KO); the protein that forms neuronal gap junctions between cortical inhibitory cells. However, genetically modified knock-out animals may exhibit compensatory effects; with the risk that observed differences between WT and Cx36 KO animals could be erroneously attributed to Cx36 gap junction effects. In this study we investigated the effect of GABA(A)-receptor modulation (augmentation with 16µM etomidate and blockade with 100µM picrotoxin) on low-magnesium seizure-like events (SLEs) in mouse cortical slices. In WT slices, picrotoxin enhanced both the amplitude (49% increase, p=0.0006) and frequency (37% increase, p=0.005) of SLEs; etomidate also enhanced SLE amplitude (18% increase, p=0.003) but reduced event frequency (25% decrease, p<0.0001). In Cx36 KO slices, the frequency effects of etomidate and picrotoxin were preserved, but the amplitude responses were abolished. Pre-treatment with the gap junction blocker mefloquin in WT slices did not significantly alter the drug responses, indicating that the reduction in amplitude seen in the Cx36 KO mice was not primarily mediated by their lack of interneuronal gap junctions, but was rather due to pre-existing compensatory changes in these animals. Conclusions from studies comparing seizure characteristics between WT and Cx36 KO mice must be viewed with a degree of caution because of the possible confounding effect of compensatory neurophysiological changes in the genetically modified animals.

Role of Cx36 gap junction modulation in general anaesthetic anticonvulsant action.

Many GABAergic anaesthetics reduce gap junction coupling but it is currently unknown whether this effect contributes to anaesthetic anticonvulsant action. In this study we examined the possible role of connexin36 gap junctions in the anticonvulsant action of isoflurane and compared this to etomidate, an anaesthetic known for having proconvulsant effects. We compared the effect of anaesthetic concentrations of isoflurane (1 MAC) and etomidate (16 microM) on low-magnesium-induced interictal-like activity in isolated neocortical slices. The effect of connexin36 gap junction blockade was explored by comparing effects in slices from wild-type mice and from a transgenic mouse strain lacking the gene for connexin36. In slices from wild-type mice, both isoflurane (1 MAC) and etomidate (16 microM) reduced interictal-like event frequency; mean(S.D.) reduction of 44(13)% (P<0.0001) and 25(24)% (P<0.0001), respectively. The reduction in event frequency was greater for isoflurane (P<0.005). Isoflurane had no effect on the amplitude of interictal-like events, but event amplitude was enhanced by etomidate (18(28)% increase, P<0.005). The capacity for isoflurane to reduce event frequency was significantly reduced, but not eliminated in slices from connexin36 knock-out mice (33(15)% reduction, P<0.05 for the difference with wild-type), while that of etomidate remained unchanged (23(39)% reduction). The etomidate-mediated increase in event amplitude was eliminated in connexin36 knock-out slices. The results from this study support the hypothesis that the anticonvulsant effect of isoflurane is in part mediated by gap junction blockade. The role of gap junction modulation by etomidate is more complicated and may be important in the mechanism of action of etomidate's proconvulsant effects.