Degenerative myelopathy in 18 Pembroke Welsh Corgi dogs.

Postmortem examination was performed on 18 Pembroke Welsh Corgi dogs (mean age 12.7 years) with clinical signs and antemortem diagnostic tests compatible with a diagnosis of degenerative myelopathy. Tissue sections from specific spinal cord and brain regions were systematically evaluated in all dogs. Axonal degeneration and loss were graded according to severity and subsequently compared across different spinal cord segments and funiculi. White matter lesions were identified in defined regions of the dorsal, lateral, and ventral funiculi. The dorsolateral portion of the lateral funiculus was the most severely affected region in all cord segments. Spinal cord segment T12 exhibited the most severe axonal loss. Spinal nerve roots, peripheral nerves, and brain sections were within normal limits, with the exception of areas of mild astrogliosis in gray matter of the caudal medulla. Dogs with more severe lesions showed significant progression of axonal degeneration and loss at T12 and at cord segments cranial and caudal to T12. Severity of axonal loss in individual dogs positively correlated with the duration of clinical signs. The distribution of axonal degeneration resembled that reported in German Shepherd Dog degenerative myelopathy but differed with respect to the transverse and longitudinal extent of the lesions within more clearly defined funicular areas. Although these lesion differences might reflect disease longevity, they could also indicate a form of degenerative myelopathy unique to the Pembroke Welsh Corgi dog.

Pharmacokinetics and toxicity of bromide following high-dose oral potassium bromide administration in healthy Beagles.

The pharmacokinetics of a multidose regimen of potassium bromide (KBr) administration in normal dogs was examined. KBr was administered at 30 mg/kg p.o. q 12 h for a period of 115 days. Serum, urine, and cerebrospinal fluid (CSF) bromide (BR) concentrations were measured at the onset of dosing, during the accumulation phase, at steady-state, and after a subsequent dose adjustment. Median elimination half-life and steady-state serum concentration were 15.2 days and 245 mg/dL, respectively. Apparent total body clearance was 16.4 mL/day/kg and volume of distribution was 0.40 L/kg. The CSF:serum BR ratio at steady-state was 0.77. Dogs showed no neurologic deficits during maintenance dosing but significant latency shifts in waves I and V of the brainstem auditory evoked response were evident. Following a subsequent dose adjustment, serum BR concentrations of approximately 400 mg/dL were associated with caudal paresis in two dogs. Estimated half-life during the accumulation phase was shorter than elimination half-lives reported in other studies and was likely related to dietary chloride content. The range of steady-state concentrations achieved suggests individual differences in clearance and bioavailability between dogs. The described protocol reliably produced serum BR concentrations that are required by many epileptic patients for satisfactory seizure control.

Evolution of a molecular switch: universal bacterial GTPases regulate ribosome function.

The GTPases comprise a protein superfamily of highly conserved molecular switches adapted to many diverse functions. These proteins are found in all domains of life and often perform essential roles in fundamental cellular processes. Analysis of data from genome sequencing projects demonstrates that bacteria possess a core of 11 universally conserved GTPases (elongation factor G and Tu, initiation factor 2, LepA, Era, Obg, ThdF/TrmE, Ffh, FtsY, EngA and YchF). Investigations aimed at understanding the function of GTPases indicate that a second conserved feature of these proteins is that they elicit their function through interaction with RNA and/or ribosomes. An emerging concept suggests that the 11 universal GTPases are either necessary for ribosome function or transmitting information from the ribosome to downstream targets for the purpose of generating specific cellular responses. Furthermore, it is suggested that progenitor GTPases were early regulators of RNA function and may have existed in precursors of cellular systems driven by catalytic RNA. If this is the case, then a corollary of this hypothesis is that GTPases that do not bind RNA arose at a later time from an RNA-binding progenitor that lost the capability to bind RNA.

Treatment of partial seizures and seizure-like activity with felbamate in six dogs.

Six dogs with partial seizures or partial seizure-like activity were treated with the antiepileptic drug felbamate between 1993 and 1998. All dogs had a history and results of diagnostic testing suggestive of either primary (idiopathic) or occult secondary epilepsy. Dogs ranged between four months and eight years of age at the onset of seizure activity. The median time period between onset of the first seizure and the start of felbamate therapy was 3.8 months (range 0.75 to 36 months). Median duration of therapy was nine months (range two to 22 months). All dogs experienced a reduction in seizure frequency after felbamate administration. Median total number of seizures post-treatment was two (range 0 to 9). Two dogs had an immediate and prolonged cessation of seizure activity. Steady-state trough serum felbamate concentrations measured at two weeks, and one, 12 and 22 months after the commencement of therapy in four dogs ranged between 13 and 55 mg/litre (median 35 mg/litre). Reversible haematological adverse effects were detected in two dogs, with one dog developing concurrent keratoconjunctivitis sicca. These results suggest that felbamate can be an effective antiepileptic drug without life-threatening complications when used as monotherapy for partial seizures in the dog.

Muscular dystrophy in female dogs.

The most common form of muscular dystrophy in dogs and humans is caused by mutations in the dystrophin gene. The dystrophin gene is located on the X chromosome, and, therefore, disease-causing mutations in dystrophin occur most often in males. Therefore, females with dystrophin deficiency or other forms of muscular dystrophy may be undiagnosed or misdiagnosed. Immunohistochemistry was used to analyze dystrophin and a number of other muscle proteins associated with muscular dystrophy in humans, including sarcoglycans and laminin alpha2, in muscle biopsy specimens from 5 female dogs with pathologic changes consistent with muscular dystrophy. The female dogs were presented with a variety of clinical signs including generalized weakness, muscle wasting, tremors, exercise intolerance, gait abnormalities, and limb deformity. Serum creatine kinase activity was variably high. One dog had no detectable dystrophin in the muscle; another was mosaic, with some fibers normal and others partly dystrophin-deficient. A 3rd dog had normal dystrophin but no detectable laminin alpha2. Two dogs could not be classified. This study demonstrates the occurrence of dystrophin- and laminin alpha2-associated muscular dystrophy and the difficulty in clinical diagnosis of these disorders in female dogs.

The feline model of neuroAIDS: understanding the progression towards AIDS dementia.

Feline immunodeficiency virus (FIV) is a neurotropic lentivirus that produces a protracted state of immunodeficiency and encephalopathy in the cat. Recent evidence has shown several similarities to the natural progression of human immunodeficiency virus infection (HIV-1) associated degenerative effects on the central and peripheral nervous systems. Similar to HIV-1, FIV-induced encephalopathy neurovirulence is strain dependent, results in progressive immunodeficiency and increasing early peripheral but not brain viral load, preferentially affects the developing nervous system, produces quantifiable behavioural and neurophysiological impairment that is not directly linked to neuronal infectivity, and induces neuronal injury and loss both in vivo and in vitro. This paper highlights the cumulative scientific body of evidence supporting the use of the feline model of neuroAIDS.

Alterations in the peptidyltransferase and decoding domains of ribosomal RNA suppress mutations in the elongation factor G gene.

The translocation stage of protein synthesis is a highly conserved process in all cells. Although the components necessary for translocation have been delineated, the mechanism of this activity has not been well defined. Ribosome movement on template mRNA must allow for displacement of tRNA-mRNA complexes from the ribosomal A to P sites and P to E sites, while ensuring rigid maintenance of the correct reading frame. In Escherichia coli, translocation of the ribosome is promoted by elongation factor G (EF-G). To examine the role of EF-G and rRNA in translocation we have characterized mutations in rRNA genes that can suppress a temperature-sensitive (ts) allele of fusA, the gene in E. coli that encodes EF-G. This analysis was performed using the ts E. coli strain PEM100, which contains a point mutation within fusA. The ts phenotype of PEM100 can be suppressed by either of two mutations in the decoding region of the 16S rRNA when present in combination with a mutation at position 2058 in the peptidyltransferase domain of the 23S rRNA. Communication between these ribosomal domains is essential for coordinating the events of the elongation cycle. We propose a model in which EF-G promotes translocation by modulating this communication, thereby increasing the efficiency of this fundamental process.

Highly efficient, enantioselective synthesis of (+)-grandisol from a C2-symmetric bis(alpha,beta-butenolide).

[reaction: see text] A new, very efficient, enantioselective synthesis of the sexual attracting insect pheromone (+)-grandisol has been developed, in which the key step is the double [2 + 2] photocycloaddition of ethylene to a bis(alpha,beta-butenolide) readily available from D-mannitol. The C2 symmetry of the substrate and the appropriate protection of the central diol unit are the crucial features for the high diastereofacial discrimination during the cycloaddition process.

Acute bilateral trigeminal neuropathy associated with nervous system lymphosarcoma in a dog.

A nine-year-old dog presented with clinical signs consistent with bilateral trigeminal neuropathy. Multicentric lymphoma was diagnosed, and neoplastic lymphocytes were identified in the cerebrospinal fluid. Electromyography revealed spontaneous activity in temporal and masseter muscles. Histopathological examination demonstrated neoplastic cell invasion of temporal and masseter myofibers and of multiple peripheral nerves, including the trigeminal nerve. Central nervous system pathology consisted primarily of spinal root and leptomeningeal lymphoid cell infiltration with relative sparing of spinal cord and brain parenchyma.

A biological assay for detection of heterogeneities in the surface hydrophobicity of polymer coatings exposed to the marine environment.

Minimally adhesive polymers are being developed as potential coatings for use in the marine environment. A 'bioprobe', the bacterium Psychrobacter sp. strain SW5, was employed to detect heterogeneities in substratum hydrophobicity at a micrometer level, rather than the millimeter level detected by traditional contact angle measurements. This novel assay was based on substratum-induced shifts in bacterial morphology and was used to demonstrate that characteristics of these surfaces can be evaluated for maintenance of parameters such as low surface free energy as well as temporal stability when immersed in water. Immersion of developmental substrata in artificial seawater for up to 90d prior to testing with the bioprobe potentially affects the stability of the designed characteristics of the polymers. It is proposed that the shifts in cell and biofilm morphology results from changes influencing the surface hydrophobicity of the polymers. An unpredicted outcome of this testing was the detection of modifications to coatings inferred by the addition of filler particles. Exposure of coatings to the natural microbial community of seawater revealed colonization characteristics that substantiate the results obtained by using the bioindicator.

The function of conserved amino acid residues adjacent to the effector domain in elongation factor G.

Bacterial elongation factor G (EF-G) physically associates with translocation-competent ribosomes and facilitates transition to the subsequent codon through the coordinate binding and hydrolysis of GTP. In order to investigate the amino acid positions necessary for EF-G functions, a series of mutations were constructed in the EF-G structural gene (fusA) of Escherichia coli, specifically at positions flanking the effector domain. A mutated allele was isolated in which the wild-type sequence from codons 29 to 47 ("EFG2947") was replaced with a sequence encoding 28 amino acids from ribosomal protein S7. This mutated gene was unable to complement a fusAts strain when supplied in trans at the nonpermissive temperature. In vitro biochemical analysis demonstrated that nucleotide crosslinking was unaffected in EFG2947, while ribosome binding appeared to be completely abolished. A series of point mutations created within this region, encoding L30A, Y32A, H37A, and K38A were shown to give rise to fully functional proteins, suggesting that side chains of these individual residues are not essential for EF-G function. A sixth mutant, E41A, was found to inefficiently rescue growth in a fusAts background, and was also unable to bind ribosomes normally in vitro. In contrast E41Q could restore growth at the nonpermissive temperature. These results can be explained within the context of a three-dimensional model for the effector region of EF-G. This model indicates that the effector domain contains a negative potential field that may be important for ribosome binding.

An imposed oscillating electrical field improves the recovery of function in neurologically complete paraplegic dogs.

We show that an applied electric field in which the polarity is reversed every 15 minutes can improve the outcome from severe, acute spinal cord injury in dogs. This study utilized naturally injured, neurologically complete paraplegic dogs as a model for human spinal cord injury. The recovery of paraplegic dogs treated with oscillating electric field stimulation (OFS) (approximately 500 to 600 microV/mm; n = 20) was compared with that of sham-treated animals (n = 14). Active and sham stimulators were fabricated in West Lafayette, Indiana. They were coded, randomized, sterilized, and packaged in Warsaw, Indiana, and returned to Purdue University for blinded surgical implantation. The stimulators were of a previously unpublished design and meet the requirements for phase I human clinical testing. All dogs were treated within 18 days of the onset of paraplegia. During the experimental applications, all received the highest standard of conventional management, including surgical decompression, spinal stabilization (if required), and acute administration of methylprednisolone sodium succinate. A radiologic and neurologic examination was performed on every dog entering the study, the latter consisting of standard reflex testing, urologic tests, urodynamic testing, tests for deep and superficial pain appreciation, proprioceptive placing of the hind limbs, ambulation, and evoked potential testing. Dogs were evaluated before and after surgery and at 6 weeks and 6 months after surgery. A greater proportion of experimentally treated dogs than of sham-treated animals showed improvement in every category of functional evaluation at both the 6-week and 6-month recheck, with no reverse trend. Statistical significance was not reached in comparisons of some individual categories of functional evaluation between sham-treated and OFS-treated dogs (ambulation, proprioceptive placing); an early trend towards significance was shown in others (deep pain), and significance was reached in evaluations of superficial pain appreciation. An average of all individual scores for all categories of blinded behavioral evaluation (combined neurologic score) was used to compare group outcomes. At the 6-month recheck period, the combined neurologic score of OFS-treated dogs was significantly better than that of control dogs (p = 0.047; Mann-Whitney, two-tailed).

Effects of feline immunodeficiency virus on cognition and behavioral function in cats.

Experimental intravenous challenge of 8-week old cats with the Maryland isolate of feline immunodeficiency virus, Maryland isolate (FIV-MD) was investigated for its effects on cognitive and behavioral function at 12 months postinfection. Six cats infected with FIV-MD were compared with age-matched controls on several behavioral measures. These measures included an open field observation, locomotion tests, traversing planks of various widths for food reinforcement, and a spatial learning task. No group differences were observed on any measure of locomotion. Differences were present with exploratory and stationary activity in the open field observation, with infected cats exhibiting higher levels of exploratory activity and in less stationary activity compared with that of control cats. In the plank-walking experiment, infected cats were less able to successfully cross progressively narrower planks compared with control animals. A holeboard paradigm was constructed to test spatial learning and memory, in which cats were required to locate food reinforcement based on position in the holeboard array. As a group, FIV-infected cats committed more reference (exploring an unbaited cup) and working memory (returning to a previously visited baited cup) errors than control cats. The main difference demonstrated was a higher activity level and associated distractibility in FIV-infected cats that appears to be related to their overall deficient performance when learning new tasks. These results indicate that behavioral function is altered and cognition is quantitatively impaired in FIV-infected cats.

Evaluation of hypothermia-induced analgesia and influence of opioid antagonists in leopard frogs (Rana pipiens).

Hypothermia results in diminished voluntary muscle activity, and is frequently used as a means of providing deep anesthesia to ectotherms and some mammals. In ectotherms, however, it is unclear if hypothermia produces true pain insensation. A needle-probe thermometer was used to demonstrate in frogs (Rana pipiens) that local hypothermia (9 degrees C) could be induced by placement of a tourniqueted leg into ice water (6 degrees C) for 10 min in contrast to the contralateral nontourniqueted leg (21.8 degrees C) kept out of ice water. Analgesia was tested by placement of dilutions of acetic acid on the rear leg. Further tests using groups of 10 frogs demonstrated that frogs with local hypothermia tolerated greater concentrations of acetic acid (mean acetic acid test score = 11) than morphine (10 mg/kg)-treated (9.6) or nontreated (5.8) frogs. Additional studies showed that morphine analgesia was blocked with naloxone doses as low as 0.01 mg/kg and hypothermia-induced analgesia at 10 mg/kg. Naltrexone blocked morphine analgesia at dosages as low as 0.01 mg/kg and hypothermia-induced analgesia at 0.10 mg/kg. In summary, this study demonstrates that hypothermia induces significant analgesia in an amphibian, and that this analgesia is partially blocked by naloxone and naltrexone, suggesting that the effect is mediated at least partially by opioid receptors.

Seizures: classification, etiologies, and pathophysiology.

Epilepsy is a disorder of recurrent seizures that are neural in origin. Partial seizures are usually due to a structural cerebrocortical lesion and may be simple or complex. Brain injuries may alter inherent neuronal properties and neuronal circuits and lead to recurrent excitatory activity. Potentiation of excitatory synapses and depression of inhibitory synapses are probable critical events in epileptogenesis. The pathogenic factors underlying primary or idiopathic generalized seizures are not as well understood. A more diffuse or multifocal state of neuronal excitability may be the result of early congenital events that are magnified over time. The progression of subclinical neuronal excitatory activity to a clinical seizure may relate directly to the phenomenon of cortical plasticity.

Molecular genetics of bacterial attachment and biofouling.

Microbial adhesion to animate or inert surfaces is potentially mediated by nonspecific physical or specific ligand-receptor interactions. Growth and survival of the microbial community or biofilm then depends on adaptation to a series of changing environmental milieux. Within the realm of cell-cell interaction, recent advances suggest that flagella, fimbriae and other protein receptors are essential for bacterial attachment to surfaces. There has also been profound progress in the elucidation of genes and molecules necessary for bacterial attachments to surfaces and subsequent biofilm formation.

GABAergic neuroaxonal dystrophy and other cytopathological alterations in feline Niemann-Pick disease type C.

Feline Niemann-Pick disease type C (NPC) is an autosomal recessive lysosomal storage disease which shares many of the clinical, biochemical and pathological features of the corresponding human disorder. Cytopathological alterations in distinct neuronal cell populations were investigated in this animal model to gain a better understanding of the pathogenesis of brain dysfunction. Golgi and immunocytochemical methods were employed to characterize the cell architectural changes occurring in neuronal somata, dendrites and axons at different stages of disease progression. Cortical pyramidal neurons in laminae II, III, and V exhibited various degrees of meganeurite and/or swollen axon hillock formation with or without ectopic dendritogenesis. Enlarged axon hillock regions with neuritic processes and spines were recognized early in the progression of feline NPC but were less prevalent in mid to late stages of the disease. Glutamic acid decarboxylase (GAD) immunocytochemistry demonstrated immunoreactive spheroids in numerous GABAergic axons in neocortex, subcortical areas, and cerebellum. Parvalbumin-immunoreactive axonal spheroid distribution in brain closely mirrored results from the GAD studies, whereas calbindin D-28k-immunoreactive spheroids were conspicuously absent in most cortical and subcortical areas examined. Purkinje cell axonal spheroid formation progressed in a distal to proximal direction, with eventual involvement of recurrent axon collaterals. Purkinje cell death and a concomitant decrease in the numbers of spheroids in the cerebellum were observed late in the disease course. Clinical neurological signs in feline NPC occur in parallel with neuronal structural alterations and suggest that GABAergic neuroaxonal dystrophy is a contributor to brain dysfunction in this disease.