Anticonvulsant drugs and the genetically epilepsy-prone rat

Anticonvulsant drugs were evaluated in members of two colonies of genetically epilepsy-prone rats (GEPR). Virtually all of the animals in the first colony experience a wild running fit that terminates in a generalized clonic convulsion when they are stimulated by sound. According to our convulsion intensity scoring system, these animals have an audiogenic response score (ARS) of 3 and the colony is designated the GEPR-3 colony. In the second colony, more than 95% of the animals experience a wild running phase terminating in a tonic extensor convulsion when they are stimulated by sound. That is, they have an ARS of 9 and the colony is designated the GEPR-9 colony. All of the established antiepileptic drugs that were tested produced anticonvulsant effects in the GEPR. Three tricyclic antidepressant agents acted as anticonvulsants in doses substantially lower than the toxic doses that produced spontaneous convulsions. Two of the established anticonvulsants, phenobarbital and ethosuximide, produced anticonvulsant effects in very similar doses in members of GEPR-3 and GEPR-9 colonies. Valproic acid produced an anticonvulsant effect in GEPR-3 in significantly lower doses than in GEPR-9. Carbamazepine, phenytoin, imipramine, amitriptyline, and desipramine produced anticonvulsant effects in essentially equimolar doses and in each case the protective dose was significantly lower in GEPR-9 than in GEPR-3 colonies. GEPR did not experience the convulsive effects of imipramine, amitriptyline, and desipramine at lower doses than did control animals. Thus, these epilepsy-prone animals are no more likely to experience convulsions in response to overdose of one of these three drugs than are nonepileptic subjects.     

Personal digital assistant (PDA) cell phone units produce elevated extremely-low frequency electromagnetic field emissions

Initial tests indicate that personal and occupational use of personal digital assistants (PDAs or palm-held wireless units) produce high intensity bursts of extremely-low frequency electromagnetic fields (ELF-EMF). These emissions could result in comparatively high ELF-EMF exposure in persons that carry a PDA close to the body (i.e., in a pocket or on a belt); or held to the head for cell phone conversations. ELF-EMF emissions of 10 microT were recorded on PDAs during normal office use over a 24 h test period. Results of ELF-EMF measurements show that email transmit and receive functions produce rapid, short-duration ELF-EMF spikes in the 2-10 microT range, each lasting several seconds to over a minute apparently depending on file download size. Some units produced spikes as high as 30-60 microT during email activities. Cell phone activity on PDAs produced continuously elevated ELF-EMF readings in the 0.5-1 microT range, as opposed to the rapid spiking pattern for email receipt and transmission. Switching the PDA unit from "OFF" to "ON" position resulted in single ELF-EMF pulses of over 90 microT on two units. Email downloads into the PDA can occur randomly throughout the day and night when the unit is "ON"; thus the user who wears the PDA may be receiving high-intensity ELF-EMF pulses throughout the day and night. The frequency of email traffic on the PDA, and the power switching unit (battery unit) may affect the frequency and intensity of ELF-EMF emissions.     

Mixed self-assembly of polydiacetylenes for highly specific and sensitive strip biosensors

Polydiacetylenes (PDAs), which possess unique properties that allow them to change color in response to environmental changes such as variations in pH, temperature, and molecular binding, have been widely investigated as signal transducers in biosensor applications. Most PDA-based sensors reported to date have been evaluated largely on the basis of their ability to detect purified samples, however, and their specificity has rarely been tested. In this study, novel PDAs fabricated on polyvinylidene fluoride (PVDF) strips by photoreaction of composite diacetylene self-assemblies were developed as biosensors, and nonspecific binding to off-target biomolecules was assessed. A mixed PDA surface containing biotin and ethanolamide bound the target, i.e., streptavidin, more specifically than did biotin alone. The optimized PDA biosensor exhibited approximately 2850-fold higher selectivity for streptavidin relative to bovine serum albumin controls. A PDA biosensor that was further prepared showed distinctive signals for the urine of diabetic patients compared to urine samples from healthy/non-diabetic person due to the concentration of microalbuminuria. To our knowledge, this is the first strip-type biosensor fabricated with PDAs and the first PDA-based biosensor that can effectively overcome the problem of nonspecific binding.     

The genetically epilepsy-prone rat: an overview of seizure-prone characteristics and responsiveness to anticonvulsant drugs

The Genetically Epilepsy-Prone Rat (GEPR) is rapidly gaining support as a model of epilepsy. In addition to a marked sensitivity to both sound-induced and hyperthermic seizures, GEPRs exhibit unusual sensitivity to a number of seizure-provoking modalities, including various forms of electrical and chemical stimulation. The existence of a moderate seizure colony (GEPR-3) and a severe seizure colony (GEPR-9) allows pathophysiological studies of seizure susceptibility and severity. The consistency of seizures within each colony allows for comparisons in seizure naive GEPRs and seizure experienced GEPRs. The consistent seizure responses of the GEPR are also ideal for the testing of anticonvulsant drugs. Further, the relative potencies of anticonvulsant drugs between the two colonies of GEPRs predict the clinical efficacies of traditional antiepileptic drugs and may be able to predict novel anticonvulsants.     

Sleep, temperature, activity, and prolactin phenotypes of genetically epilepsy-prone rats

Sleep-wake disturbances are common in epilepsy, yet the potential adverse effect of seizures on sleep is not well characterized. Genetically epilepsy-prone rats (GEPRs) are a well-studied model of genetic susceptibility to audiogenic seizures. To assess their suitability for investigating relationships between seizures and disordered sleep, we characterized the sleep, activity, and tempera ture patterns of 2 GEPR strains (designated 3 and 9) and Sprague-Dawley (SD) rats in the basal state, after forced wakefulness, and after exposure to sound-induced seizures at light onset and dark onset. Because of observed differences in rapid-eye-movement sleep (REMS), we also assessed serum levels of prolactin, which is implicated in REMS regulation. The data reveal that under basal conditions, the GEPR3 strain shows less SWS and REMS, higher core temperatures, and higher serum prolactin concentrations than do GEPR9 and SD strains. All 3 strains respond similarly to enforced sleep loss. Seizures induced at light onset delay the onset of SWS in both GEPR strains. Seizures induced at dark onset do not significantly alter sleep. Genotype assessment indicates that although both GEPR strains are inbred (that is, homozygous at 107 genetic markers), they differ from each other at 74 of 107 loci. Differences in basal sleep, temperature, and prolactin between GEPR3 and GEPR9 strains suggest different homeostatic regulation of these functions. Our detection of concurrent alterations in sleep, temperature, and prolactin in these 2 GEPR strains implicates the hypothalamus as a likely site for anatomic or physiologic variation in the control of these homeostatic processes.     

Patent ductus arteriosus in adults

Patent ductus arteriosus (PDA) is a congenital heart defect in which the ductus arteriosus, a vascular structure between the pulmonary artery and the aorta that normally closes shortly after birth, remains open. We present two cases of adults with PDA. A 28-year-old lady had a small PDA without any symptoms or left heart overload. The PDA was closed for endocarditis prevention using a coil. In a 37-year-old lady with palpitations, collapse, dyspnoea and left heart overload caused by a large PDA, percutaneous closure of the PDA was performed with an Amplatzer device. Transcatheter closure has been established to be the method of choice for treating a PDA in adults. (Neth Heart J 2008;16:255-9.)     

The architectural design of networks of protein domain architectures

Protein domain architectures (PDAs), in which single domains are linked to form multiple-domain proteins, are a major molecular form used by evolution for the diversification of protein functions. However, the design principles of PDAs remain largely uninvestigated. In this study, we constructed networks to connect domain architectures that had grown out from the same single domain for every single domain in the Pfam-A database and found that there are three main distinctive types of these networks, which suggests that evolution can exploit PDAs in three different ways. Further analysis showed that these three different types of PDA networks are each adopted by different types of protein domains, although many networks exhibit the characteristics of more than one of the three types. Our results shed light on nature''s blueprint for protein architecture and provide a framework for understanding architectural design from a network perspective.     

Patent ductus arteriosus in adults – indications and possibilities for closure

Patent ductus arteriosus (PDA) is a rare diagnosis in adults, since symptoms and signs usually occur in infancy and most cases are treated shortly after diagnosis. We present two patients who were first diagnosed with PDA during adulthood. The first case represents a more severe form of PDA, where the need for closure of the PDA is obvious. In the second case the sequelae of the PDA are less clear. In both patients, closure of the PDA (surgically in one case, percutaneously in the other) was successful.     

Cerebral cortical GABA and benzodiazepine binding sites in genetically seizure prone rats

Adult male and female genetically seizure-prone rats were assessed for sound-induced seizures. Heterozygous control groups were compared with mild seizure (designated GEPR 3) and severe seizure animals (GEPR 9). Groups of animals were killed and crude synaptosome fractions (P2) prepared from freshly dissected cerebral cortices. Binding sites for gamma-aminobutyric acid (GABA) were assessed by [3H]-muscimol in the absence or presence of excess GABA and/or pentobarbital. Binding sites for benzodiazepines were assessed by [3H]-flunitrazepam in the presence or absence of clonazepam. Compared to controls, GEPR 3 animals had a modest increase and GEPR 9 animals a larger increase in Bmax for both high and low affinity GABA sites, with no change in Kd. Chloride-dependent, barbiturate-enhanced GABA binding (increased Bmax) was observed in all conditions and groups. Likewise benzodiazepine binding (Bmax) increased slightly in GEPR 9 animals. There were no observed changes in binding sites for a survey of biogenic amines. Seizure-prone animals appear to have compensatory denervation-like supersensitivity for their most prominent inhibitory receptor, which may or may not be linked to the seizure event.     

Altered plasma proteins released from platelets and endothelial cells are associated with human patent ductus arteriosus

Patent ductus arteriosus is the third most common congenital heart disease and resulted from the persistence of ductal patency after birth. Ductus arteriosus closure involves functional and structural remodeling, controlled by many factors. The changes in plasma protein levels associated with PDA closure are not known. Here we for the first time demonstrate six key differential plasma proteins in human patent ductus arteriosus patients using proteomic technology and present a model to illustrate the constriction and closure of ductus arteriosus. Differentially expressed proteins were analyzed by using isobaric tags for relative and absolute quantification and validated by enzyme-linked immunosorbent assay in new samples. The proteomic data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the data set identifier PXD008568. We found 74 upregulated and 98 downregulated proteins in the plasma of patients with PDA. Five decreased proteins (platelet factor 4, fibrinogen, von Willebrand factor, collagen, and mannose binding lectin-associated serine protease-2) and one increased protein (fibronectin) may increase the risk of patent ductus arteriosus. Those proteins are closely related to platelet activation and coagulation cascades, complement mannan-binding-lectin, and other systemic signaling pathways. Our findings for the first time indicate that the differential proteins involved in different pathways may play key roles in the nonclosure of the ductus arteriosus in humans and may be developed as biomarkers for diagnosis. All those findings may be served as the basis of understanding the etiology and pathogenesis of patent ductus arteriosus.     

Elevation of naloxone-sensitive 3H-dihydromorphine binding in hippocampal formation of genetically epilepsy-prone rats

3H-Dihydromorphine (DHM) binding sites were measured in the brain of non-epileptic control and GEPR rats using in vitro autoradiographic techniques. The number of naloxone-sensitive 3H-DHM binding sites was increased 38-57% in the pyramidal cell layer of ventral hippocampal CA3 and Ca1 of GEPR-3 and GEPR-9 rats compared to non-epileptic controls. No significant differences in 3H-DHM binding were observed in dorsal hippocampal formation, lateral entorhinal cortex, lateral geniculate or cerebellum. The results suggest that an increase in the number of opioid receptors in ventral hippocampus of GEPR rats may be one factor contributing to the enhanced sensitivity of GEPR-9 rats to the proconvulsant effects of morphine.     

Anatomical changes of the GABAergic system in the inferior colliculus of the genetically epilepsy-prone rat

The number of GABAergic neurons as determined by GAD immunocytochemistry and total neurons as determined from Nissl preparations were counted and classified at the light microscopic level in the inferior colliculus (IC) of the genetically epilepsy prone rat (GEPR) and the non-epileptic Sprague-Dawley (SD) strain of rat. GAD-positive neurons are abundant in the IC and a significant increase in the number of GAD-positive neurons occurs in the GEPR as compared to the SD in all three subdivisions. However, the most pronounced difference occurs in the ventral lateral portion of the central nucleus, where there is a selective increase in the small (200%) and medium-sized (90%) GABAergic somata (10-15 microns in diameter and 15-25 microns in diameter, respectively). As determined from Nissl preparations an increase in total numbers of neurons also occurs. Thus, a 100% increase in the number of small neurons and a 30% increase in the number of medium-sized neurons occur in the adult GEPR as compared to the SD rat. A statistically significant increase in the numbers of small neurons also occurred in the IC of the young GEPR. At 4 days of age, a 55% increase in the number of small neurons was found, and at 10 days of age this increase was 105%. The numbers of the medium and large neurons were similar in the older group of rats. These data suggest that the increase in cell number observed in the adult GEPR is not compensatory to the seizure activity, but may either be genetically programmed or be a failure of cell death. Based on other studies of genetic models of epilepsy, we propose that the additional GABAergic neurons may disinhibit excitatory projection neurons in the IC.     

The development of kindled seizures is accelerated in the genetically epilepsy-prone rat

The kindling phenomenon was examined in genetically epilepsy-prone (GEPR) and non-epileptic control Sprague-Dawley rats. Kindling stimulations were administered three times a day until each rat had exhibited three Class 5 kindled motor seizures. The mean total number of kindling stimulations required for each experimental group to exhibit three motor seizures of each motor seizure class was determined. The results indicated that the early stage of kindling development was accelerated significantly in both the GEPR-3 and GEPR-9 rats, compared to non-epileptic control rats. Later stages of kindling development were accelerated in GEPR-9 but not GEPR-3 rats. Thus a differential acceleration of kindling development was exhibited by GEPR-3 and GEPR-9 rats. The results suggest the possibility that some brain region(s) involved in the early stages of kindling development may be hyperexcitable in both GEPR-3 and GEPR-9 rats. Other brain region(s) involved with the later stages of kindling development may be more excitable in GEPR-9 rats. These putative alterations may, in part, contribute to the seizure prone state of GEPR rats and the differential seizure responses of GEPR-3 and GEPR-9 rats.     

Vesicular stomatitis virus as an oncolytic agent against pancreatic ductal adenocarcinoma

Vesicular stomatitis virus (VSV) is a promising oncolytic agent against a variety of cancers. However, it has never been tested in any pancreatic cancer model. Pancreatic ductal adenocarcinoma (PDA) is the most common and aggressive form of pancreatic cancer. In this study, the oncolytic potentials of several VSV variants were analyzed in a panel of 13 clinically relevant human PDA cell lines and compared to conditionally replicative adenoviruses (CRAds), Sendai virus and respiratory syncytial virus. VSV variants showed oncolytic abilities superior to those of other viruses, and some cell lines that exhibited resistance to other viruses were successfully killed by VSV. However, PDA cells were highly heterogeneous in their susceptibility to virus-induced oncolysis, and several cell lines were resistant to all tested viruses. Resistant cells showed low levels of very early VSV RNA synthesis, indicating possible defects at initial stages of infection. In addition, unlike permissive PDA cell lines, most of the resistant cell lines were able to both produce and respond to interferon, suggesting that intact type I interferon responses contributed to their resistance phenotype. Four cell lines that varied in their permissiveness to VSV-ΔM51 and CRAd dl1520 were tested in mice, and the in vivo results closely mimicked those in vitro. While our results demonstrate that VSV is a promising oncolytic agent against PDA, further studies are needed to better understand the molecular mechanisms of resistance of some PDAs to oncolytic virotherapy.     

Patent ductus arteriosus in mice with smooth muscle-specific Jag1 deletion

The ductus arteriosus is an arterial vessel that shunts blood flow away from the lungs during fetal life, but normally occludes after birth to establish the adult circulation pattern. Failure of the ductus arteriosus to close after birth is termed patent ductus arteriosus and is one of the most common congenital heart defects. Mice with smooth muscle cell-specific deletion of Jag1, which encodes a Notch ligand, die postnatally from patent ductus arteriosus. These mice exhibit defects in contractile smooth muscle cell differentiation in the vascular wall of the ductus arteriosus and adjacent descending aorta. These defects arise through an inability to propagate the JAG1-Notch signal via lateral induction throughout the width of the vascular wall. Both heterotypic endothelial smooth muscle cell interactions and homotypic vascular smooth muscle cell interactions are required for normal patterning and differentiation of the ductus arteriosus and adjacent descending aorta. This new model for a common congenital heart defect provides novel insights into the genetic programs that underlie ductus arteriosus development and closure.     

Paternal Age and Offspring Congenital Heart Defects: A National Cohort Study

Paternal age has been associated with offspring congenital heart defects (CHDs), which might be caused by increased mutations in the germ cell line because of cumulated cell replications. Empirical evidences, however, remain inconclusive. Furthermore, it is unknown whether all subtypes of CHDs are affected by paternal age. We aimed to explore the relationship between paternal age and the risk of offspring CHDs and its five common subtypes using national register data in Denmark. A total of 1 893 899 singletons born in Denmark from 1977 to 2008 were included in this national-based cohort study. Cox’s proportion hazards model with robust sandwich estimate option was used to estimate the hazards ratio (95% confidence interval) for the associations between paternal age and all CHDs, as well as subtypes of CHDs (patent ductus arteriosus (PDA), ventricular septal defect (VSD), atrial septal defect (ASD), tetralogy of fallot (TOF) and coarctation of the aorta (CoA)). We did not observe an overall association between paternal age and offspring CHDs. However, compared to the paternal age of 25–29 years, paternal age of older than 45 years was associated with a 69% increased risk of PDA (HR45+ = 1.69, 95%CI:1.17–2.43). We observed similar results when subanalyses were restricted to children born to mothers of 27–30 years old. After taking into consideration of maternal age, our data suggested that advanced paternal age was associated with an increased prevalence of one subtype of offspring congenital heart defects (CHDs), namely patent ductus arteriosus (PDA).     

Neurotransmitter abnormalities in genetically epileptic rodents

A growing body of evidence supports a pathophysiological role for norepinephrine (NE) and serotonin in the regulation of seizures in the genetically epilepsy-prone rat (GEPR). Other evidence indicates that gamma-aminobutyric acid (GABA) and taurine may also participate in the seizure regulation process. Innate deficits in NE and serotonin appear to be causes of the genetically determined seizure-prone states of the GEPR, whereas abnormalities in GABAergic systems and taurine metabolism may represent inadequate attempts of the central nervous system to compensate for the seizure-prone state in these rats. In audiogenic seizure-susceptible (AGS) mice, evidence suggests a role for dopamine as well as GABA and possibly serotonin. NE may contribute to the regulation of seizures in AGS mice, but consistent evidence for a primary role for this monoamine is lacking. It is suggested that there is no single common neurotransmitter abnormality underlying genetic seizure disorders in humans or other animals and that the GEPR and the AGS mouse may both serve as good models for study of the neurochemical abnormalities that underlie the different human epilepsies.     

Noradrenergic and serotonergic determinants of seizure susceptibility and severity in genetically epilepsy-prone rats

Pharmacological studies demonstrate a reciprocal relationship between both noradrenergic and serotonergic transmission and audiogenic seizure severity and susceptibility in the genetically epilepsy-prone rat (GEPR). In contrast, drug-induced changes in the neurochemical indices of dopaminergic activity do not result in alterations in seizure severity. These pharmacological investigations led to the hypothesis that both noradrenergic and serotonergic neurons are capable of regulating seizure severity in the GEPR. Pharmacological investigations also provided evidence that monoaminergic neurons serve as determinants of seizure susceptibility in these epileptic animals. The GEPR is susceptible to environmentally-induced seizures which cannot be precipitated in neurologically normal subjects. Drug studies suggest that monoaminergic decrements serve as one set of susceptibility determinants. However, non-monoaminergic abnormalities also play important roles in the seizure predisposition which characterizes the GEPR. Pathophysiological studies have confirmed and extended the concepts generated by the pharmacological investigations. Noradrenergic and serotonergic deficits do indeed characterize the seizure naive state of the GEPR. These studies have provided a basis for tentative identification of areas of the brain in which monoaminergic abnormalities regulate seizure severity and susceptibility. Monoaminergic defects in some areas such as the thalamus may regulate both susceptibility and severity. In other areas, defects may regulate only severity or susceptibility. In the striatum, noradrenergic defects do not appear to be present and probably are not determinants of the epileptic state of the GEPR.