(R)-[11C]Verapamil PET studies to assess changes in P-glycoprotein expression and functionality in rat blood-brain barrier after exposure to kainate-induced status epilepticus

<p>Abstract</p> <p>Background</p> <p>Increased functionality of efflux transporters at the blood-brain barrier may contribute to decreased drug concentrations at the target site in CNS diseases like epilepsy. In the rat, pharmacoresistant epilepsy can be mimicked by inducing status epilepticus by intraperitoneal injection of kainate, which leads to development of spontaneous seizures after 3 weeks to 3 months. The aim of this study was to investigate potential changes in P-glycoprotein (P-gp) expression and functionality at an early stage after induction of status epilepticus by kainate.</p> <p>Methods</p> <p><it>(R)</it>-[¹¹C]verapamil, which is currently the most frequently used positron emission tomography (PET) ligand for determining P-gp functionality at the blood-brain barrier, was used in kainate and saline (control) treated rats, at 7 days after treatment. To investigate the effect of P-gp on <it>(R)</it>-[¹¹C]verapamil brain distribution, both groups were studied without or with co-administration of the P-gp inhibitor tariquidar. P-gp expression was determined using immunohistochemistry in post mortem brains. <it>(R)</it>-[¹¹C]verapamil kinetics were analyzed with approaches common in PET research (Logan analysis, and compartmental modelling of individual profiles) as well as by population mixed effects modelling (NONMEM).</p> <p>Results</p> <p>All data analysis approaches indicated only modest differences in brain distribution of <it>(R)</it>-[¹¹C]verapamil between saline and kainate treated rats, while tariquidar treatment in both groups resulted in a more than 10-fold increase. NONMEM provided most precise parameter estimates. P-gp expression was found to be similar for kainate and saline treated rats.</p> <p>Conclusions</p> <p>P-gp expression and functionality does not seem to change at early stage after induction of anticipated pharmacoresistant epilepsy by kainate.</p

Nestin-GFP Transgene Reveals Neural Precursor Cells in Adult Skeletal Muscle

Background: Therapy for neural lesions or degenerative diseases relies mainly on finding transplantable active precursor cells. Identifying them in peripheral tissues accessible for biopsy, outside the central nervous system, would circumvent the serious immunological and ethical concerns impeding cell therapy. Methodology/Principal Findings: In this study, we isolated neural progenitor cells in cultured adult skeletal muscle from transgenic mice in which nestin regulatory elements control GFP expression. These cells also expressed the early neural marker Tuj1 and light and heavy neurofilament but not S100b, indicating that they express typical neural but not Schwann cell markers. GFP+/Tuj1+ cells were also negative for the endothelial and pericyte markers CD31 and a-smooth muscle actin, respectively. We established their a) functional response to glutamate in patch-clamp recordings; b) interstitial mesenchymal origin; c) replicative capacity; and d) the environment necessary for their survival after fluorescenceactivated cell sorting. Conclusions/Significance: We propose that the decline in nestin-GFP expression in muscle progenitor cells and its persistence in neural precursor cells in muscle cultures provide an invaluable tool for isolating a population of predifferentiated neural cells with therapeutic potential

Multilocus ISSR Markers Reveal Two Major Genetic Groups in Spanish and South African Populations of the Grapevine Fungal Pathogen Cadophora luteo-olivacea

Cadophora luteo-olivacea is a lesser-known fungal trunk pathogen of grapevine which has been recently isolated from vines showing decline symptoms in grape growing regions worldwide. In this study, 80 C. luteo-olivacea isolates (65 from Spain and 15 from South Africa) were studied. Inter-simple-sequence repeat-polymerase chain reaction (ISSR-PCR) generated 55 polymorphic loci from four ISSR primers selected from an initial screen of 13 ISSR primers. The ISSR markers revealed 40 multilocus genotypes (MLGs) in the global population. Minimum spanning network analysis showed that the MLGs from South Africa clustered around the most frequent genotype, while the genotypes from Spain were distributed all across the network. Principal component analysis and dendrograms based on genetic distance and bootstrapping identified two highly differentiated genetic clusters in the Spanish and South African C. luteo-olivacea populations, with no intermediate genotypes between these clusters. Movement within the Spanish provinces may have occurred repeatedly given the frequent retrieval of the same genotype in distant locations. The results obtained in this study provide new insights into the population genetic structure of C. luteo-olivacea in Spain and highlights the need to produce healthy and quality planting material in grapevine nurseries to avoid the spread of this fungus throughout different grape growing regions

Targeting prostaglandin E2 EP1 receptors prevents seizure-associated P-glycoprotein up-regulation

Up-regulation of the blood-brain barrier efflux transporter P-glycoprotein in central nervous system disorders results in restricted brain access and limited efficacy of therapeutic drugs. In epilepsies, seizure activity strongly triggers expression of P-glycoprotein. Here, we identified the prostaglandin E2 receptor, EP1, as a key factor in the signaling pathway that mediates seizure-induced up-regulation of P-glycoprotein at the blood-brain barrier. In the rat pilocarpine model, status epilepticus significantly increased P-glycoprotein expression by 92 to 197% in the hippocampal hilus and granule cell layer as well as the piriform cortex. The EP1 receptor antagonist 8-chlorodibenz[b,f][1,4]oxazepine-10(11H)-carboxylic acid, 2-[1-oxo-3-(4- pyridinyl)propyl]hydrazide hydrochloride (SC-51089) abolished seizure-induced P-glycoprotein up-regulation and retained its expression at the control level. The control of P-glycoprotein expression despite prolonged seizure activity suggests that EP1 receptor antagonism will also improve antiepileptic drug efficacy. Preliminary evidence for this concept has been obtained using a massive kindling paradigm during which animals received a subchronic SC-51089 treatment. After withdrawal of the EP1 receptor antagonist, a low dose of the P-glycoprotein substrate phenobarbital resulted in an anticonvulsant effect in this pretreated group, whereas the same dosage of phenobarbital did not exert a significant effect in the respective control group. In conclusion, our data demonstrate that EP1 is a key signaling factor in the regulatory pathway that drives P-glycoprotein up-regulation during seizures. These findings suggest new intriguing possibilities to prevent and interrupt P-glycoprotein overexpression in epilepsy. Future studies are necessary to further evaluate the appropriateness of the strategy to enhance the efficacy of antiepileptic drugs

Targeting prostaglandin E2 EP1 receptors prevents seizure-associated P-glycoprotein up-regulation

Up-regulation of the blood-brain barrier efflux transporter P-glycoprotein in central nervous system disorders results in restricted brain access and limited efficacy of therapeutic drugs. In epilepsies, seizure activity strongly triggers expression of P-glycoprotein. Here, we identified the prostaglandin E2 receptor, EP1, as a key factor in the signaling pathway that mediates seizure-induced up-regulation of P-glycoprotein at the blood-brain barrier. In the rat pilocarpine model, status epilepticus significantly increased P-glycoprotein expression by 92 to 197% in the hippocampal hilus and granule cell layer as well as the piriform cortex. The EP1 receptor antagonist 8-chlorodibenz[b,f][1,4]oxazepine-10(11H)-carboxylic acid, 2-[1-oxo-3-(4-pyridinyl)propyl]hydrazide hydrochloride (SC-51089) abolished seizure-induced P-glycoprotein up-regulation and retained its expression at the control level. The control of P-glycoprotein expression despite prolonged seizure activity suggests that EP1 receptor antagonism will also improve antiepileptic drug efficacy. Preliminary evidence for this concept has been obtained using a massive kindling paradigm during which animals received a subchronic SC-51089 treatment. After withdrawal of the EP1 receptor antagonist, a low dose of the P-glycoprotein substrate phenobarbital resulted in an anticonvulsant effect in this pretreated group, whereas the same dosage of phenobarbital did not exert a significant effect in the respective control group. In conclusion, our data demonstrate that EP1 is a key signaling factor in the regulatory pathway that drives P-glycoprotein up-regulation during seizures. These findings suggest new intriguing possibilities to prevent and interrupt P-glycoprotein overexpression in epilepsy. Future studies are necessary to further evaluate the appropriateness of the strategy to enhance the efficacy of antiepileptic drugs