Isocard.org

Van Amstel S.R. et al./ Journal of Camelid Science 2 (2009) 41-49 Ivermectin concentrations in serum and cerebrospinal fluid after
intravenous administration to healthy Llamas
Van Amstel S. R.*, Portmann A. B., Doherty T. J, Newman S. J, Yarbrough J. W and
College of Veterinary Medicine, University of Tennessee, 2407 River drive, Knoxville, Tennessee 37996.
Abstract
The response to anthelmintic treatment in clinical cases of meningeal worm (Parelaphostrongylus tenuis) infestation remains unpredictable. In a previous study, ivermectin (IVM) was not detected in cerebrospinal fluid (C.S.F.) following subcutaneous administration to healthy llamas of IVM at 500µg/kg. In the present study, the same IVM dose was added to 1 L 0.9% NaCl and administered intravenously over 30 minutes to 6 healthy llamas. C.S.F and blood were collected, at baseline and 2, 4, 6, 12, 24, 48 and 60 hr after IVM administration. Serum and C.S.F. were stored at -400C and IVM concentration was determined using high performance liquid chromatography (H.P.L.C.). No IVM was detected in the serum or C.S.F. in baseline samples. Concentrations of IVM ranging from 2291- 7742 ng/ml were present in serum at 2 hr post dosing but values decreased to between 103-615 ng/ml at the 4 hr sampling, and ranged from 11- 48 ng/ml at 60hr. Low concentrations of IVM were found in the C.S.F. of all llamas on at least one of the sampling times. Immediately after administration of IVM three of the llamas showed transient lethargy and decreased appetite for 12-24 hours. One llama developed acute neurological signs 7 days after IVM administration and was euthanased three days later. Histopathologic examination revealed diffuse myelinic oedema in the brain and spinal cord. Two other llamas developed C.S.F. changes consistent with septic inflammation. C.S.F. concentrations of IVM can be achieved following I.V. administration at 500µg/kg, however, because of the possibility of neurological damage, I.V. administration of IVM at this dose is not recommended. Key words: blood, cerebrospinal fluid, ivermectin concentrations, llama.


1. Introduction
and chronic multifocal microinfarctions. Intralesional parasites can be demonstrated microscopically. Larvae may also migrate for meningeal worm (Parelaphostrongylus tenuis) and do not become clinically affected. Adult worms occur in the veins and sinuses of the dura mater and cause no the treatment of clinical cases of meningeal injury to the brain or spinal cord. Camelids goats) on the other hand are aberrant hosts. lactones produced by fermentation of the spinal nerves to the spinal cord where they (Reinemeyer, 2001). It is highly lipophilic Van Amstel S.R. et al./ Journal of Camelid Science 2 (2009) 41-49 metabolites are avermectin B1 derivatives with fecal excretion as the main route of administration to llamas (Lama glama) and anthelminitic, such as ivermectin, with the (<2-8ng/ml) and which may take up to 8 potential to penetrate into the brain and spinal cord should be efficacious in cases 2004, Oukessou et al., 1996, Oukessou et of P. tenuis infestation; however, the al., 1999, Jarnivinen et al., 2002). A report on sub cutaneous (S.C.) administration of complicates the therapeutic use of IVM in peak serum concentrations of 3ng/ml after 7days (Jarvinen et al., 2002). In another penetration through the blood brain barrier study, IVM was not detected in the blood nervous system depends to a large extent multidrug efflux transporter P-glycoprotein 2003). Serum concentrations in the other 5 (P.G.P.) in the endothelial cells of the animals ranged from 1.4 – 16.4 ng/ml (Van blood brain barrier. P.G.P. is responsible study, no IVM was detected in the C.S.F. inside cells back into the blood (Kwei et al., 1999, Schinkel et al., 1994, Tamai and neurological signs, low concentrations (2.9 C.S.F. perhaps as a result of changes in the endothelial capillaries (Nobmann et al., whether detectable concentrations of IVM can be obtained in the C.S.F. of healthy ivermectin present in the central nervous system leading to neurological signs by its administration of the drug at 500µg/kg. action as a γ-aminobutyric acid agonist, 2. Materials and Methods
Anecdotally, IVM (200 – 500 µg/kg) is subcutaneously to llamas with neurological disease suspected to be due to P. tenuis infestation, however the efficacy of IVM is 2. 1. Llamas
study. The llamas were housed indoors in Van Amstel S.R. et al./ Journal of Camelid Science 2 (2009) 41-49 2.3 Sampling Schedule
were collected (1 & 5 ml, respectively) at Health, Ford Dodge, Iowa, 50501) 3 weeks baseline (24 hr after catheter placement) prior to the start of the experiment and and 2, 4, 6, 12, 24, 48 and 60hr after IVM vaccinated with Clostridium perfringens types C and D and Clostridium tetani stored at –40 0C until IVM analysis. Spinal following collection of the 60hr sample. A 2.2 Catheterization and ivermectin
evaluated cytologically to determine the administration
Jugular and spinal catheters were placed 2.4 Ivermectin Analysis
guidewire style, 14ga x 20cm) was placed spinal fluid (C.S.F.) samples and spinal cord tissue in case of the one animal that technique and flushed every 4 hours with (AnaSed® injection, Lloyd Laboratories, fluorescence detector (Waters Corporation, Health, Ford Dodge, Iowa, 50501, 2mg/kg) Epidural Anesthesia Set, B Braun Medical mobile phase was an isocratic mixture of fluid flowed freely through the needle, the methanol, acetonitrile, and water (56:40:4). catheter was inserted and advanced about 3 All solutions were prepared using double- distilled, deionized water filtered (0.22 catheter was then secured to the skin just distal to the point of entry. The calculated rate was 1.7 ml/min and the fluorescence IVM (Ivomec® 1% injection for cattle and detector was set to an excitation of 365 nm and vortexed. One milliliter of sample was placed in a 7 ml screw cap test tube and mixed with 25 µl internal standard (moxidectin 1 µg/ml), and 1 ml mixture of acetonitrile:water (80:20) then placed on a Van Amstel S.R. et al./ Journal of Camelid Science 2 (2009) 41-49 tube rocker for 20 minutes. Samples were ng/ml for serum samples, and 0.1 to 1000 Spiked standards were treated exactly as solid phase extraction cartridge (Waters 64% for the spinal cord sample. Intra-assay N2 in a 47oC water bath. Sample residues were derived using 100 µ l imidazole: while inter-assay variability ranged from trifluoroacetic anhydride: acetonitrile (1:2). 3. Results
placed into H.P.L.C. vials, and a 20 µl reported here due to the inability to obtain Preparation of spinal tissue was done by C.S.F. from the remaining llama. Three of weighing out one gram of spinal tissue into the llamas (# 2, 4 & 5) showed transient of acetonitrile water (80:20) was added to the tube and the mixture was homogenized depression; drooping of the lower lip; lip using a Fisher Scientific brand PowerGen and ear twitching; sternal recumbency with the head down and the neck stretched out; respiratory rates were noted. The clinical signs and changes in behavior lasted for 12 adverse signs following I.V. administration of IVM and remained bright and alert with normal food intake during the whole trial methanol was evaporated under nitrogen in acute neurological signs seven days after with 100 l of imidazole : Acetonitrile (1:1) and then 150l trifluoroacetic acid anhydride : acetonitrile (1:2). The solution rigidity in all four legs. The llama was able seconds and then loaded into an HPLC vial to rise unaided shortly afterwards but was ataxic, and hypermetric. Twenty four hours later the llama was in sternal recumbency, and spinal tissue analyses were produced unable to rise and developed torticollis. linear concentration ranges of 1 to 9000 Van Amstel S.R. et al./ Journal of Camelid Science 2 (2009) 41-49 vertebral spinal cord and cerebellar coning thoraco-lumbar spine and this finding was with blunting of cerebral gyri, indicative of thought to rule out traumatic injury. The being multifocal involving the brainstem localised to the central nervous system and and forebrain. Results of a complete blood more specifically to the white matter of the entire spinal cord, with lesser involvement were unremarkable except for the presence in the white matter tracts of the brain. of a stress leukogram and hyperglycemia. Mild to moderate distension of the myelin C.S.F. analysis indicated a non-suppurative mixed inflammation, which was attributed which had been removed 4 days earlier at cervical and lumbar spinal cord sections Table 1. Mean concentration and range of
ivermectin in the serum and C.S.F. of five llamas given ivermectin at 500 µg /kg. astrocytes were characterised by abundant Time post
injection
Serum IVM
diagnosis for the brain and spinal cord was noted histologically would have accounted for the cerebellar herniation and cerebral gyri blunting noted grossly. represented a single incident rather than multiple insults and, based on the degree of a number of llamas sampled = 4; b number of llamas oligodendroglial cell death, were relatively acute in onset (within 3-5 days of death). 3.2 Serum ivermectin concentrations
euthanased on the third day after having samples. High concentrations of IVM were present in the 2hr samples, ranging from Van Amstel S.R. et al./ Journal of Camelid Science 2 (2009) 41-49
Table 2. Concentrations of ivermectin (ng/ml) in serum and C.S.F. of individual llamas at specific
sampling times.
ND = none detected; NS = sample not procured ivermectin
102 – 615 ng/ml with a mean of 491 ng/ml. concentrations
Thereafter, IVM concentrations decreased samples with final serum concentrations at 3.5 Cytological examination of C.S.F.
3.3 Cerebrospinal fluid ivermectin
concentrations
IVM concentration in the C.S.F. was found Van Amstel S.R. et al./ Journal of Camelid Science 2 (2009) 41-49
4. Discussion
concentrations were detected in the C.S.F. depression, drooping of the lower lip, lip This amounted to an average of 0.25 ng/ml down and the neck stretched out, decreased administration may not be the cause of this C.S.F. of 8 healthy llamas 24 hours after reaction but the vehicle propylene glycol three consecutive S.C. injections of IVM may be implicated because vehicle-treated 24 hours apart at a dose rate of 500µg/kg the conclusion that IVM is very effectively 2001). One llama in the study (#1 Table 2) developed neurological signs 7 days after I.V. administration despite the fact that IVM was only detected in the C.S.F. at the concentrations decreased by approximately 4hr sampling Table 2. However samples of 90% between the 2 and 4 hr samples after the spinal cord was positive for ivermectin 50-60% for each of the follow up samples. rate). This is similar to a study by Seaman et al., (1987) who found an average of 56 on a) A wide therapeutic index: In general, ug/kg avermectin B1a in brain and spinal IVM has at least a tenfold safety margin in cord tissue from 5 calves showing clinical ruminants (Reinemeyer, 2001). In cattle, signs of toxicity. In the same study spinal toxic effects generally do not appear until cord from one affected steer contained 34 animal (Seaman et al., 1987). The delay in developed ataxia, listless and occasionally onset of neurological signs in the present death. Sheep showed ataxia and depression IVM has been used intravenously in sheep were not reported in that study (Canga et having received IVM intramuscularly at a al., 2007). b) Poor absorption following dose rate of 330 µg/kg, two animals died S.C. administration: No IVM was detected injections of I.V.M. 24 hours apart at a penetration of the blood brain barrier by IVM as a result of the action of drug efflux Van Amstel S.R. et al./ Journal of Camelid Science 2 (2009) 41-49 al.,1987). However in one report on IVM in calves ( Seaman et al., 1987, Button et toxicity in a group of dogs belonging to the collie breed onset of signs varied from 3 Nevertheless, although administration of be achieved following I.V. administration IVM could potentially have had neurotoxic whether such low levels would be effective administration to the onset of neurologic signs remains perplexing. The neurological the possibility of neurological damage, I.V. reported for other animals including calves administration of IVM at this dose is not Clinical signs in a group of calves included “suddenly falling down” which is exactly References
what was first seen in this llama (Button et Burkholder T.H., Jensen J., Chen H., Junkins K., al., 1988). Other frequently reported signs Chatfield J., Boothe D., 2004. Plasma evaluation for ivermectin in llamas (lama of IVM toxicity also seen in the llama in glama) after standard subcutaneous dosing. J this study include ataxia and hypermetria, Button C., Barton R., Honey P., Rickford P., 1988. activity and comatose state (Seaman et al., Avermectin toxicity in calves and an evaluation 1987, Button et al., 1988). The myelinic of picrotoxin as an antidote. Aust Vet J., 65(5),157-158. Canga A.G., Sahagun A., Diez M.J., Fernandez N., Sierra M., Garcia J.J., 2007. Bioavailability of a cytotoxic oedema that is often the result of commercial formulation of ivermectin after osmotic dysregulation, either locally or subcutaneous administration to sheep. AJVR., Edwards G., 2003. Ivermectin: does P-glycoprotein play a role in neurotoxicity? Filaria J., 2 (Suppl Jarvinen J.A., Miller J.A., Oehler D.D., 2002. abnormalities and particularly sodium and Pharmacokinetics of ivermectinin llamas (Lama Kwei G.Y., Alvaro R.F., Chen Q., Jenkins H.J., parameters were considered within normal Hop C.E.A.C., Keohane V.T.L.Y., Strauss J.R., limits for this llama. Cytotoxic oedema can also occur following nutritional deficiency, ischaemia/hypoxia or exposure to toxicants ivermectin and cyclosporin A in CF-1 mice such as IVM (Summers et al., 1995, Maxie deficient in MDR1A p-glycoprotein. Drug Metab Dispos., 27(5),581-587. Maxie M.G, Youssef S. 2007. Nervous system. In:. Pathology of Domestic Animals5th ed. Maxie MG., (Ed) Saunders Elsevier, Toronto Canada Ivermectin excretion by isolated functionally intact brain endothelial capillaries. Br J histopathological lesions in the brain or spinal cord could be demonstrated in both Oukessou M., Badri M., Sutra J.F., Galtier P., accidental and experimental IVM toxicity Van Amstel S.R. et al./ Journal of Camelid Science 2 (2009) 41-49 ivermectin in the camel (Camelus dromedarius). Summers B.A., Cummings T.F., DeLahunta A., 1995. Principles of Neuropathology. Chapter 1. Oukessou M., Berrag B., Alvinerie M., 1999. A In: Veterinary Neuropathology. Summers BA comparative kinetic study of ivermectin and Tamai I., Tsuji A., 2000. Transporter-mediated dromedaries). Vet Parasitol., 83, 151-159. permeation of drugs across the blood-brain Plumb D.C., 2005. Ivermectin. In: Plumb’s barrier. J Pharm Sci., 89(11), 1371-1381 Veterinary Drug Handbook., Plumb D.C., (Ed). Summers B.A., Cummings T.F., DeLahunta A., Stockholm, Wisconsin., Pharmavet Inc., 435. 1995. Inflammation of the nervous system Reinemeyer C.R., 2001. Chemotherapy of parasitic Chapter 3. In: Veterinary Neuropathology. diseases. In: Veterinary Pharmacology and Summers BA (Ed). Mosby St Louis, Mo publ., Therapeutics. Adams HR (Ed). Iowa State Press Van Amstel SR, Miller AJ. Ivermectin levels in Schinkel A.H., Smit J.J.M., van Tellingen O., blood and cerebrospinal fluid from healthy Beijnin J.H., Wagenaar E., van Deemter L., Mol llamas and cases with neurological signs. Proc of the 21st annual ACVIM Forum, Charlotte, Maandag E.C., te Riele H.P.J., Berns A.J.M., North Carolina, USA, 4-8 June. 2003, 944-945. Borst P., 1994. Disruption of the mouse mdr1a Van der Lugt J.J, Venter I. 2007. Myelin p-glycoprotein gene leads to a deficiency in the vacuolation, optic neuropathy and retinal blood-brain barrier and to increased sensitivity degeneration after closantel overdosage in sheep Seaman J.T, Eagleson J.S, Carrigan M.J., Webb R.F., 1987. Avermectin B1 toxicity in a herd of Murray Grey cattle. Aust Vet J., 64(9), 284-285.

Source: http://www.isocard.org/e_Library/Journal_of_Camelid_Science/JCS_2009_Volume_02/06_Ivermectin.pdf