Changes in Retinal Function and Morphology Are Early Clinical Signs of Disease in Cattle with Bovine Spongiform Encephalopathy

Bovine spongiform encephalopathy (BSE) belongs to a group of fatal, transmissible protein misfolding diseases known as transmissible spongiform encephalopathies (TSEs). All TSEs are caused by accumulation of misfolded prion protein (PrPSc) throughout the central nervous system (CNS), which results in neuronal loss and ultimately death. Like other protein misfolding diseases including Parkinson’s disease and Alzheimer’s disease, TSEs are generally not diagnosed until the onset of disease after the appearance of unequivocal clinical signs. As such, identification of the earliest clinical signs of disease may facilitate diagnosis. The retina is the most accessible part of the central nervous system, and retinal pathology in TSE affected animals has been previously reported. Here we describe antemortem changes in retinal function and morphology that are detectable in BSE inoculated animals several months (up to 11 months) prior to the appearance of any other signs of clinical disease. We also demonstrate that differences in the severity of these clinical signs reflect the amount of PrPSc accumulation in the retina and the resulting inflammatory response of the tissue. These results are the earliest reported clinical signs associated with TSE infection and provide a basis for understanding the pathology and evaluating therapeutic interventions

Evaluation of a combinatorial approach to prion inactivation using an oxidizing agent, SDS, and proteinase K

BACKGROUND: Prions demonstrate an unusual resistance to methods effective at inactivating conventional microorganisms. This has resulted in a very tangible and difficult infection control challenge to the medical and veterinary communities, as well as animal agriculture and related industries. Currently accepted practices of harsh chemical treatments such as prolonged exposure to sodium hydroxide or sodium hypochlorite, or autoclaving are not suitable in many situations. Less caustic and more readily applicable treatments to contaminated environments are therefore desirable. We recently demonstrated that exposure of the RML scrapie agent to a commercial product containing sodium percarbonate (SPC-P) with or without sodium dodecyl sulfate (SDS) rendered PrP(Sc) sensitive to proteinase K (PK), but did not eliminate infectivity. The current study was designed to evaluate the efficacy of a combinatorial approach to inactivating prions by exposing RML-positive brain homogenate to SPC-P and SDS followed by PK. Treated samples were evaluated for PrP(Sc)-immunoreactivity by western blot, and residual infectivity by mouse bioassay. RESULTS: Treatment of infected brain homogenate with SPC-P and SDS followed by PK exposure resulted in a 4–5 log(10) reduction in infectivity when bioassayed in tga20 mice. CONCLUSIONS: This study demonstrates that exposure of the RML scrapie agent to SPC-P and SDS followed by PK markedly reduces, but does not eliminate infectivity. The results of this study encourage further investigation into whether consecutive or concomitant exposure to sodium percarbonate, SDS, and a protease may serve as a viable and non-caustic option for prion inactivation

Temporal Resolution of Misfolded Prion Protein Transport, Accumulation, Glial Activation, and Neuronal Death in the Retinas of Mice Inoculated with Scrapie

Currently, there is a lack of pathological landmarks to describe the progression of prion disease in vivo. Our goal was to use an experimental model to determine the temporal relationship between the transport of misfolded prion protein (PrPSc) from the brain to the retina, the accumulation of PrPSc in the retina, the response of the surrounding retinal tissue, and loss of neurons. Retinal samples from mice inoculated with RML scrapie were collected at 30, 60, 90, 105, and 120 days post inoculation (dpi) or at the onset of clinical signs of disease (153 dpi). Retinal homogenates were tested for prion seeding activity. Antibody staining was used to assess accumulation of PrPSc and the resulting response of retinal tissue. Loss of photoreceptors was used as a measure of neuronal death. PrPSc seeding activity was first detected in all samples at 60 dpi. Accumulation of PrPSc and coincident activation of retinal glia were first detected at 90 dpi. Activation of microglia was first detected at 105 dpi, but neuronal death was not detectable until 120 dpi. Our results demonstrate that by using the retina we can resolve the temporal separation between several key events in the pathogenesis of prion disease

Altered electroretinogram b-wave in a Suffolk sheep experimentally infected with scrapie

TRANSMISSIBLE spongiform encephalopathies (TSEs) are a group of fatal neurodegenerative diseases in which an abnormal isoform of the cellular prion protein (PrPSc) accumulates in tissues of the central nervous system. Accumulation of PrPSc occurs in the retina, a rostral projection of the central nervous system, of both natural and nonnatural host species with TSEs (Foster and others 1999, Spraker and others 2002, Head and others 2003, 2005, Hamir and others 2004, 2005, Kercher and others 2004, Greenlee and others 2006, Hortells and others 2006). In retinas from scrapie-affected sheep, PrPSc accumulation is primarily observed in the inner and outer plexiform layers, and in the ganglion cell layer (Jeffrey and others 2001, Greenlee and others 2006). Recent studies have reported few (Hortells and others 2006) or no (Greenlee and others 2006) histological lesions in the retinas of sheep affected with scrapie. However, morphological changes in specific retinal cell types have been demonstrated (Smith and others 2008). Despite the morphological consequences of retinal PrPSc accumulation in sheep (Barnett and Palmer 1971, Smith and others 2008), the functional impact on the retina of these animals is unknown. In the current study, the effect of TSE on retinal function in a scrapie-infected Suffolk sheep using flash electroretinography was investigated

Assessment Techniques Corresponding to Scientific Texts in Commercial Reading Programs: Do They Promote Scientific Literacy?

This research is part of a larger study of commercial reading programs used in Canada in grades 1-6. The specific purposes of the results reported here were to identify and quantify the assessment techniques suggested for the selections that contain scientific content, to show how the assessments differ by grade, to evaluate the nature and quality of the assessments, and to examine the extent to which the assessments help foster scientific literacy. It was found that the assessments occurred in six major forms and employed about a dozen assessment tools that engage students in nearly 20 tasks. Such variety is endorsed in both literacy and science education position statements. The assessments showed some weak trends by grade, but primarily left the purpose of the assessments to teachers’ judgment. The consequence is that teachers probably will choose the assessments for formative rather than summative evaluation, an approach also endorsed by literacy and science education policy statements. Hardly any of the assessments focused on the specificities of learning to read texts that are scientific such as interpreting descriptions of methods and research findings and thus had limited use in promoting this particular aspect of scientific literacy

A medium throughput rodent model of relapse from addiction with behavioral and pharmacological specificity

One of most formidable problems in the treatment of addiction is the high rate of relapse. The discovery of medicines to help mitigate relapse are aided by animal models that currently involve weeks of training and require surgical preparations and drug delivery devices. The present set of experiments was initiated to investigate a rapid 8-day screening method that utilizes food instead of intravenous drug administration. Male Sprague-Dawley rats were trained in a reinstatement paradigm in which every lever press produced a 45 mg food pellet concurrently paired with a light and tone. Behavior was subsequently extinguished with lever responses producing neither food nor food-associated stimuli. Reinstatement of responding was evaluated under conditions in which the first three responses of every 5 min time bin produced a food pellet along with food-associated stimuli. The mGlu5 receptor antagonists MPEP and MTEP produced a significant reduction in reinstatement while failing to alter responding where every response produced food. The cannabinoid CB1 receptor antagonist rimonabant and the mGlu2/3 receptor agonist LY379268 also selectively reduced reinstatement. Other compounds including clozapine, d-amphetamine, chlordiazepoxide, ABT-431, naltrexone and citalopram were without effect. The results suggest that relapse-like behavioral effects can be extended to non-pharmacological reinforcers. Drug effects demonstrated both behavioral and pharmacological specificity. The present experimental design thus allows for efficient and rapid assessment of the effects of drugs that might be useful in the treatment of addiction-associated relapse

Genetic variation of the prion protein gene (PRNP) in alpaca (Vicugna pacos)

Transmissible spongiform encephalopathies (TSE) are caused by accumulation of a misfolded form of the prion protein (PrP). The normal cellular isoform of PrP is produced by the prion gene (PRNP) and is highly expressed in the central nervous system. Currently, there is an absence of information regarding the genetic sequence of alpaca PRNP and the potential susceptibility of this species to TSE. The objective of this study was to sequence the open reading frame of the alpaca prion gene and analyze this sequence for variation within the alpaca population and for homology to TSE-susceptible species. We sequenced the open reading frame of the prion gene of 40 alpacas of Huacaya or Suri descent. Length polymorphisms were identified within the sampled population. A subset (15%) of animals contained an additional 24 base pairs within the putative octapeptide repeat region. This polymorphism was independent of breed and sex. The majority (52.5%) of animals were heterozygous, possessing both longer and shorter alleles. Comparison with proven TSE-susceptible species (sheep, cattle, deer) revealed the following amino acid sequence variations: I6M, A16V, M17T, G92del, Q95_G96insG, N111S, R167K, N/T177S, I206V, S225Y, Y228S, Q230G, and L237del. Sequence alignment showed high homology compared to camel (\u3e 95%), sheep (\u3e 88%), cattle (\u3e 87%) and deer (\u3e 88%) PRNP sequence. This study demonstrates intraspecies variability within the PRNP open reading frame in alpacas and overall high sequence homology to TSE-susceptible species, providing foundational data for further research on the potential susceptibility of alpacas to TSE

Retinal function and morphology are altered in cattle infected with the prion disease transmissible mink encephalopathy

Transmissible spongiform encephalopathies (TSEs) are a group of diseases that result in progressive and invariably fatal neurologic disease in both animals and humans. TSEs are characterized by the accumulation of an abnormal protease-resistant form of the prion protein in the central nervous system. Transmission of infectious TSEs is believed to occur via ingestion of prion protein–contaminated material. This material is also involved in the transmission of bovine spongiform encephalopathy (“mad cow disease”) to humans, which resulted in the variant form of Creutzfeldt-Jakob disease. Abnormal prion protein has been reported in the retina of TSE-affected cattle, but despite these observations, the specific effect of abnormal prion protein on retinal morphology and function has not been assessed. The objective of this study was to identify and characterize potential functional and morphologic abnormalities in the retinas of cattle infected with a bovine-adapted isolate of transmissible mink encephalopathy. We used electroretinography and immunohistochemistry to examine retinas from 10 noninoculated and 5 transmissible mink encephalopathy–inoculated adult Holstein steers. Here we show altered retinal function, as evidenced by prolonged implicit time of the electroretinogram b-wave, in transmissible mink encephalopathy–infected cattle before the onset of clinical illness. We also demonstrate disruption of rod bipolar cell synaptic terminals, indicated by decreased immunoreactivity for the alpha isoform of protein kinase C and vesicular glutamate transporter 1, and activation of Müller glia, as evidenced by increased glial fibrillary acidic protein and glutamine synthetase expression, in the retinas of these cattle at the time of euthanasia due to clinical deterioration. This is the first study to identify both functional and morphologic alterations in the retinas of TSE-infected cattle. Our results support future efforts to focus on the retina for the development of new strategies for the diagnosis of TSEs

Comparison of Two US Sheep Scrapie Isolates Supports Identification as Separate Strains

Scrapie is a naturally occurring transmissible spongiform encephalopathy of sheep and goats. There are different strains of sheep scrapie that are associated with unique molecular, transmission, and phenotype characteristics. However, in the United States, very little is known about the potential presence of scrapie strains. Scrapie strain and PRNP genotype could both affect susceptibility, potential for transmission, incubation period (IP), and control measures required for eliminating scrapie from a flock. The investigators evaluated 2 US scrapie isolates, No. 13-7 and x124, after intranasal inoculation to compare clinical signs, IPs, spongiform lesions, and patterns of PrPSc deposition in sheep with scrapie-susceptible PRNP genotypes (QQ171). After inoculation with x124, susceptibility and IP were associated with valine at codon 136 (V136) of the prion protein: VV136 sheep had short IPs (6.9 months), those in AV136 sheep were 11.9 months, and AA136 sheep did not develop scrapie. All No. 13-7 inoculated sheep developed scrapie, with IPs of 20.1 months for AA136 sheep, 22.8 months for AV136 sheep, and 26.7 months for VV136 sheep. Patterns of immunoreactivity in the brain were influenced by inoculum isolate and host genotype. Differences in PrPSc profiles versus isolate were most striking when examining brains from sheep with the VV136 genotype. Inoculation into C57BL/6 mice resulted in markedly different attack rates (90.5% for x124 and 5.9% for No. 13-7). Taken together, these data demonstrate that No. 13-7 and x124 represent 2 distinct strains of scrapie with different IPs, genotype susceptibilities, and PrPSc deposition profiles