Objective—To determine concentrations of excitatory
and inhibitory amino acids in CSF of a large number
of dogs with idiopathic epilepsy or genetic epilepsy
and to evaluate changes in CSF amino acid concentration
with regard to drug treatment and sex.
Animals—35 Labrador Retrievers with genetic
epilepsy (20 male and 15 female), 94 non-Labrador
Retrievers with idiopathic epilepsy (71 male and 23
female), and 20 control dogs (10 male and 10 female).
Procedure—Collection of CSF was performed > 72
hours after the occurrence of seizures. Cerebrospinal
fluid concentrations of γ-aminobutyric acid (GABA),
glutamate (GLU), aspartate (ASP), serine, and glycine
were determined by use of high performance liquid
chromatography with electrochemical detection.
Results—CSF concentrations of GABA and GLU
were significantly lower in Labrador Retrievers
with genetic epilepsy (LR-group dogs) than in control-group dogs or in non-Labrador Retrievers with
idiopathic epilepsy (non–LR-group dogs). The GLU-to-GABA ratio was significantly higher in LR-group
dogs than in non–LR-group dogs. CSF concentrations
of GLU and ASP were significantly lower
when all dogs with epilepsy (non–LR- and LR-group
dogs combined) were compared with control-group dogs.
Conclusions and Clinical Relevance—A decrease in
CSF concentrations of GABA appears to play a role in
the pathogenesis of genetically determined epilepsy
in Labrador Retrievers. However, this decrease in CSF
concentrations of GABA may also be a consequence
of seizure activity. The GLU-to-GABA ratio may prove
to be a useful indicator of genetic epilepsy in Labrador
Retrievers. (Am J Vet Res 2004;65:1108–1113)
Objective—To compare the effect of various concentrations
of sodium butyric acid and sodium valerianic
acid, as well as various osmolarities, on contractility
of ex-vivo intestinal wall specimens obtained from the
cecum and spiral colon of each of several healthy
Sample Population—Full-thickness preparations of
intestinal wall, dissected parallel to the longitudinal
smooth muscle layers, harvested from freshly slaughtered
Procedure—Specimens of intestinal wall were incubated
for 5 minutes with various concentrations of
sodium butyric acid and sodium valerianic acid as well
as various osmolar concentrations of NaCl, using a
crossover design. Isometric contractions were
induced 7 times with carbachol (CH; 5 X 10–6 mol/L).
Contractility was defined as the maximum amplitude
of contraction and the amplitude of contraction 2 minutes
after addition of CH.
Results—Repeated addition of CH did not result in a
significant effect on contractility of specimens from
the cecum and spiral colon. Contractility after addition
of CH was not significantly affected by prior incubation
with various concentrations of sodium butyric
acid or sodium valerianic acid or after an increase of
osmolarity. Maximum amplitude of contraction was
significantly higher in specimens from the spiral
colon, compared with specimens from the cecum.
Conclusions—Increases in concentrations of sodium
butyric acid or sodium valerianic acid and increases in
osmolarity did not inhibit contractility of intestinal wall
specimens from the cecum and spiral colon of a
group of healthy cows. (Am J Vet Res 2000;61:
Objective—To identify and characterize cytochrome P450 enzymes (CYPs) responsible for the metabolism of racemic ketamine in 3 mammalian species in vitro by use of chemical inhibitors and antibodies.
Sample—Human, canine, and equine liver microsomes and human single CYP3A4 and CYP2C9 and their canine orthologs.
Procedures—Chemical inhibitors selective for human CYP enzymes and anti-CYP antibodies were incubated with racemic ketamine and liver microsomes or specific CYPs. Ketamine N-demethylation to norketamine was determined via enantioselective capillary electrophoresis.
Results—The general CYP inhibitor 1-aminobenzotriazole almost completely blocked ketamine metabolism in human and canine liver microsomes but not in equine microsomes. Chemical inhibition of norketamine formation was dependent on inhibitor concentration in most circumstances. For all 3 species, inhibitors of CYP3A4, CYP2A6, CYP2C19, CYP2B6, and CYP2C9 diminished N-demethylation of ketamine. Anti-CYP3A4, anti-CYP2C9, and anti-CYP2B6 antibodies also inhibited ketamine N-demethylation. Chemical inhibition was strongest with inhibitors of CYP2A6 and CYP2C19 in canine and equine microsomes and with the CYP3A4 inhibitor in human microsomes. No significant contribution of CYP2D6 to ketamine biotransformation was observed. Although the human CYP2C9 inhibitor blocked ketamine N-demethylation completely in the canine ortholog CYP2C21, a strong inhibition was also obtained by the chemical inhibitors of CYP2C19 and CYP2B6. Ketamine N-demethylation was stereoselective in single human CYP3A4 and canine CYP2C21 enzymes.
Conclusions and Clinical Relevance—Human-specific inhibitors of CYP2A6, CYP2C19, CYP3A4, CYP2B6, and CYP2C9 diminished ketamine N-demethylation in dogs and horses. To address drug-drug interactions in these animal species, investigations with single CYPs are needed.
Objective—To describe the in vitro effects of bethanechol on contractility of smooth muscle preparations from the small intestines of healthy cows and define the muscarinic receptor subtypes involved in mediating contraction.
Sample Population—Tissue samples from the duodenum and jejunum collected immediately after slaughter of 40 healthy cows.
Procedures—Cumulative concentration-response curves were determined for the muscarinic receptor agonist bethanechol with or without prior incubation with subtype-specific receptor antagonists in an organ bath. Effects of bethanechol and antagonists and the influence of intestinal location on basal tone, maximal amplitude (Amax), and area under the curve (AUC) were evaluated.
Results—Bethanechol induced a significant, concentration-dependent increase in all preparations and variables. The effect of bethanechol was more pronounced in jejunal than in duodenal samples and in circular than in longitudinal preparations. Significant inhibition of the effects of bethanechol was observed after prior incubation with muscarinic receptor subtype M3 antagonists (more commonly for basal tone than for Amax and AUC). The M2 receptor antagonists partly inhibited the response to bethanechol, especially for basal tone. The M3 receptor antagonists were generally more potent than the M2 receptor antagonists. In a protection experiment, an M3 receptor antagonist was less potent than when used in combination with an M2 receptor antagonist. Receptor antagonists for M1 and M4 did not affect contractility variables.
Conclusions and Clinical Relevance—Bethanechol acting on muscarinic receptor sub-types M2 and M3 may be of clinical use as a prokinetic drug for motility disorders of the duodenum and jejunum in dairy cows.
Objective—To evaluate the expression of the 5-hydroxytryptamine 4 (5-HT4) receptor subtype and investigate the modulating function of those receptors on contractility in intestinal tissues obtained from horses without gastrointestinal tract disease.
Sample Population—Smooth muscle preparations from the duodenum, ileum, and pelvic flexure collected immediately after slaughter of 24 horses with no history or signs of gastrointestinal tract disease.
Procedures—In isometric organ baths, the contractile activities of smooth muscle preparations in response to 5-hydroxytryptamine and electric field stimulation were assessed; the effect of tegaserod alone or in combination with 5-hydroxytryptamine on contractility of intestinal specimens was also investigated. Presence and distribution of 5-HT4 receptors in intestinal tissues and localization on interstitial cells of Cajal were examined by use of an immunofluorescence technique.
Results—Widespread 5-HT4 receptor immunoreactivity was observed in all intestinal smooth muscle layers; 5-HT4 receptors were absent from the myenteric plexus and interstitial cells of Cajal. In electrical field–stimulated tissue preparations of duodenum and pelvic flexure, tegaserod increased the amplitude of smooth muscle contractions in a concentration-dependent manner. Preincubation with tegaserod significantly decreased the basal tone of the 5-HT–evoked contractility in small intestine specimens, compared with the effect of 5-HT alone, thereby confirming that tegaserod was acting as a partial agonist.
Conclusions and Clinical Relevance—In horses, 5-HT4 receptors on smooth muscle cells appear to be involved in the contractile response of the intestinal tract to 5-hydroxytryptamine. Results suggest that tegaserod may be useful for treatment of reduced gastrointestinal tract motility in horses.
Objective—To describe the distribution of muscarinic receptor subtypes M1 to M5 and interstitial cells of Cajal (ICCs) in the gastrointestinal tract of healthy dairy cows.
Sample Population—Full-thickness samples were collected from the fundus, corpus, and pyloric part of the abomasum and from the duodenum, ileum, cecum, proximal loop of the ascending colon, and both external loops of the spiral colon of 5 healthy dairy cows after slaughter.
Procedures—Samples were fixed in paraformaldehyde and embedded in paraffin. Muscarinic receptor subtypes and ICCs were identified by immunohistochemical analysis.
Results—Staining for M1 receptors was found in the submucosal plexus and myenteric plexus. Antibodies against M2 receptors stained nuclei of smooth muscle cells only. Evidence of M3 receptors was found in the lamina propria, in intramuscular neuronal terminals, on intermuscular nerve fibers, and on myocytes of microvessels. There was no staining for M4 receptors. Staining for M5 receptors was evident in the myocytes of microvessels and in smooth muscle cells. The ICCs were detected in the myenteric plexus and within smooth muscle layers. Distribution among locations of the bovine gastrointestinal tract did not differ for muscarinic receptor subtypes or ICCs.
Conclusions and Clinical Relevance—The broad distribution of M1, M3, M5, and ICCs in the bovine gastrointestinal tract indicated that these components are likely to play an important role in the regulation of gastrointestinal tract motility in healthy dairy cows. Muscarinic receptors and ICCs may be implicated in the pathogenesis of motility disorders, such as abomasal displacement and cecal dilatation-dislocation.
Objective—To investigate cytochrome P450 (CYP) enzymes involved in metabolism of racemic and S-ketamine in various species and to evaluate metabolic interactions of other analgesics with ketamine.
Sample Population—Human, equine, and canine liver microsomes.
Procedures—An analgesic was concurrently incubated with luminogenic substrates specific for CYP 3A4 or CYP 2C9 and liver microsomes. The luminescence signal was detected and compared with the signal for negative control samples. Ketamine and norketamine enantiomers were determined by use of capillary electrophoresis.
Results—A concentration-dependent decrease in luminescence signal was detected for ibuprofen and diclofenac in the assay for CYP 2C9 in human and equine liver microsomes but not in the assay for CYP 3A4 and methadone or xylazine in any of the species. Coincubation of methadone or xylazine with ketamine resulted in a decrease in norketamine formation in equine and canine liver microsomes but not in human liver microsomes. In all species, norketamine formation was not affected by ibuprofen, but diclofenac reduced norketamine formation in human liver microsomes. A higher rate of metabolism was detected for S-ketamine in equine liver microsomes, compared with the rate for the S-enantiomer in the racemic mixture when incubated with any of the analgesics investigated.
Conclusions and Clinical Relevance—Enzymes of the CYP 3A4 family and orthologs of CYP 2C9 were involved in ketamine metabolism in horses, dogs, and humans. Methadone and xylazine inhibited in vitro metabolism of ketamine. Therefore, higher concentrations and diminished clearance of ketamine may cause adverse effects when administered concurrently with other analgesics.