Читать книгу Large Animal Neurology - Joe Mayhew - Страница 61

The routine collection procedures for cerebrospinal fluid (CSF) sampling from large animals (Figure 3.2) have been described and simplified,^7–12 and techniques for the analysis of CSF samples have also been published.^12–19 Techniques for CSF collection from the horse are shown in Figures 3.3, 3.4, and 3.5, and those described for other large animals (Figure 3.6) can be extrapolated directly from the horse and from ruminants.^20–22 Ultrasound‐guided collection of CSF can also be considered,^23,24 and such ultrasound‐guided techniques for the collection of fluid from the atlantooccipital (Figure 3.7) and atlantoaxial spaces (Figure 3.8) in standing, adult horses can be useful in certain circumstances.^20–22,25 These are safe procedures using effective chemical restraint, although the possibility of inadvertent movement resulting in spinal cord penetration is always of concern. It is likely that worrisome signs of somnolence, stiff neck, and fever that resolves in a day or so following the cervical centesis occurs in a few cases. In neonatal foals, calves, camelids, and piglets, as well as sheep and goats, it is advantageous to use a 40–50 mm (1.5–2 inches), 20 ga disposable hypodermic needle with a plastic hub for all atlantooccipital CSF collections. These needles are exceedingly atraumatic, and in obtunded and in lightly sedated recumbent patients they cause minimal or no patient reaction. More importantly, CSF appears in the hub of the needle immediately after it enters the subarachnoid space. A sample can be obtained, and the needle withdrawn much more rapidly than if a styletted needle is used—very important in nonanesthetized patients. Indeed, during CSF collection from the cisterna magna of anesthetized dogs, it was shown that a stylet‐out technique was performed more rapidly and yielded a sample with less cellular debris than a stylet‐in technique using a standard CSF styletted needle.²⁶

Figure 3.2 Multiple tests were performed on this patient suspected of having a neuromuscular disorder. These included lumbosacral cerebrospinal fluid collection (1), needle electromyography (2), and muscle biopsies of type I and type II muscle‐fiber‐predominant muscles taken from sacrocaudalis dorsalis medialis and semitendinosus muscles at sites (indicated by 3 and 4), respectively.

Figure 3.3 Atlantooccipital cerebrospinal fluid collection from the recumbent horse. Spinal needle in position with stilette removed. Palpable landmarks are the cranial borders of the atlas (filled circles) and the external occipital protuberance (cross) on the dorsal midline.

(Source: From Mayhew¹¹ with permission.

Figure 3.4 Lumbosacral cerebrospinal fluid collection from the standing horse. Spinal needle in position with stilette removed. Palpable landmarks are the caudal borders of each tuber coxae (filled circles), the caudal edge of the spine of L₆ (star), the cranial edge of the second sacral spine (filled triangle), and the cranial edge of each tuber sacrale (filled triangle).

Source: From Mayhew¹¹ with permission.

Figure 3.5 Lumbosacral spinal fluid collection from the horse. Transverse dissection through lumbosacral articulations, cranial view. Spinal needle passes through the skin, thoracolumbar fascia adjacent to the interspinous ligaments, interarcuate ligament, dorsal dura mater and arachnoid, dorsal subarachnoid space, and conus medullaris. Needle point is in the ventral subarachnoid space. Inset shows the cranial view of pelvis, sacrum, and the area of dissection.

Source: From Mayhew¹¹ with permission.

Figure 3.6 Performing CSF collection from the atlantooccipital (AO) (A) and lumbosacral (LS) (B) sites is very straightforward with appropriate restraint, sedation, and/or general anesthesia. In very young patients light sedation is adequate, and in obtunded patients simple restraint suffices to perform an AO collection. Collection of CSF from the LS site is easier with the patient standing as the landmarks are more easily palpable and symmetric.The landmarks are shown for the AO procedure in a calf (A) with the superficial site for skin penetration indicated by . This site is at the intersection of a line joining the cranial borders of the wings of the atlas indicated by linear marks, with the midline, indicated by the spot over the occipital protuberance, while the head is flexed 90° to the neck as shown. A 19–21g, 30–45 mm hypodermic needle with plastic hub is ideal for this procedure, allowing for CSF to appear in the hub instantly the subarachnoid space is penetrated. The basic site for an LS procedure (B) is where a line joining the caudal aspects of the wings of the ilia (yellow dotted line) crosses the midline, as indicated by . This coincides with a palpable depression between the paired tubers sacrale (TS) laterally and the dorsal spines of L6 and S2 on the midline—the spine of S1 normally being too short to palpate. An 8–20 cm, 18 or 20 g spinal needle with fitted stilette is used for all adult animals depending on their size.Penetration of the terminal spinal cord during an LS procedure is essentially inconsequential. Damage to the medulla oblongata, which occurs on pathologic inspection more often than is clinically suspected, can result in additional neurologic signs.

If a patient is in an intensive care facility showing severe focal or diffuse brain injury, then consideration might be given to inserting a separate subarachnoid or subdural cranial catheter at the time of anesthesia for sampling CSF from the AO site to allow the assessment of intracranial pressure and the derivation of an estimate of cerebral perfusion pressure.^27,28 This can be of use in determining the effectiveness of ongoing management of such cases. Patient‐side observations and tests are extremely important at this stage. For example, if initial CSF is pink but clears, iatrogenic hemorrhage is likely the cause. With a sick neonatal patient, cloudy CSF with a low glucose content detected on urine test strips likely indicates suppurative meningitis. Particularly during lumbosacral sampling, small particles can often be seen to swirl in the aspirated sample. These are usually determined to be epidural fat and small aggregates of collagenous tissue introduced into the sample at the time of collection, likely reasons for falsely elevated CSF‐CK activity. When CSF analysis cannot proceed immediately following collection, the addition of a small, quantified volume (~10%) of autologous serum to an aliquot of the sample may help support cells for better cytologic harvesting, at least for 24 h.

Of note is that estimates of protein content in CSF may be substantially higher, at least when tested by the pyrogallol red method, when CSF is collected into commonly used red vacutainer tubes containing clot activator.⁽²⁹⁾

The results of CSF analysis can reflect diseases in the brain and spinal cord just as a hemogram detects many systemic disease states. Thus, CSF analysis is one of the most helpful aids in determining the nature of current and progressive CNS lesions. Normal CSF is clear and colorless with a refractive index of 1.3347–1.3350. In addition, it contains no erythrocytes and usually less than 6 × 10⁶/L (=6/μL) small mononuclear cells. Protein content is usually between 0.5 and 1.0 g/L (=50–100 mg/dL) in the horse with an absolute range of 0.1–1.2 g/L depending on the laboratory and can be up to 1.8 g/L in normal neonatal foals. Ruminants and pigs usually have CSF with a protein content of less than 0.75 g/L.¹⁸ Specific electrophoretic protein fractions in CSF can be assayed by numerous methods,¹⁷ but the utility of these results in changing the clinical course of cases is doubtful at present. Neonates of all species may have slightly xanthochromic CSF.

To help account for the blood contamination of CSF samples and to help determine the local versus systemic source of measured CSF immunoglobulin content, the determination of the albumin quotient and the IgG index, respectively, can be of some use.^30,31 Additionally, on the assumption that all red blood cells are from the circulation and not the result of previous subarachnoid bleeding, arithmetical corrections can be made for all measured components of CSF. Such corrections for routinely measured CSF components have not proven to consistently improve the utility of absolute measurements of amounts of CSF constituents.^32–34 Repeat thecal puncture from the lumbosacral or from the atlanto‐axial space 2 weeks apart has been shown not to impact results of CSF analysis.³⁵

Figure 3.7 Collection of CSF from the atlanto‐occipital cistern in a standing sedated horse. After the ears have been secured in a forward position and the area clipped and surgically prepared, a 3.5inch (8‐10 cm) 18g spinal needle is introduced and advanced as described for the procedure in recumbent anesthetized horses (A). When a distinct change in resistance (“pop”) is felt, the stylet is removed, and CSF is gently aspirated into a syringe attached to extension tubing (B). The needle may be advanced under ultrasound guidance as described by Depecker et al.²⁰

Source: Images courtesy of Carol Clark, Peterson Smith Equine Hospital & Complete Care, Ocala, FL, USA.

Many constitutive and induced enzymes have activities expressed in CSF. The cytosolic BB‐CK isoenzyme almost certainly is quite specific for neuronal tissue damage, but its measurement is somewhat complex and the interpretation of results of its content in CSF in disease states is not extremely specific. Total CK activity in CSF may well vary with disease states,³⁶ and the utility of such assays in general disease states has been questioned.³⁷ In specific neuromitochondrial disorders, lactate and pyruvate measurements may well be of diagnostic use.

Assays are available for proteins in and antibodies to infectious agents in CSF.^38,39 These tests are probably best for the subacute to chronic infections with particular viruses and protozoa. Indeed, testing for the presence of antibodies to Sarcocystis and Neospora surface antigens in CSF and blood has become popular and is very useful to assist in the diagnosis of equine protozoal myeloencephalitis,^40,41 especially when Sarcocystis neurona vaccines were used.³⁸ In particular, the ratios of specific antibody in serum to CSF are used to reveal locally produced (i.e., intrathecal) antibodies indicative of CNS infection.⁴² Unfortunately, while this antibody ratio technique is highly specific and sensitive for EPM diagnosis, it is not accurate for the diagnosis of equine neuroborreliosis.⁴³ As with many of such tests, a negative result with S. neurona immunotesting using CSF (and serum) is very useful to rule out the disease in a case with clear clinical signs in an area where the background frequency of exposure is low. However, debate arises as to interpretation of marginal test results when clinical signs are equivocal and there is a high exposure rate in the sample population. This is particularly true for a negative result, as the negative predictive value declines with increasing disease (exposure) prevalence, and for EPM the consequences of a false‐negative result (failure to treat) are more serious than a false‐positive result (unnecessary treatment). Again, with a high rate of false‐positive and false‐negative results occurring with borreliosis testing, there can be no reliance on serology for the confirmation of that disease.

One aspect of CSF chemistry that may well become useful in future is the analysis of levels of neurotransmitters, neurohormonal metabolic products, and antibodies directed against constitutive proteins to help categorize some disease states.^44–49 This will most likely relate to those diseases that have no recognized morbid neuropathologic basis, such as shivers, headshaking, narcolepsy, cataplexy, self‐mutilation, rage syndrome, acquired tremor syndromes, spastic syndrome, and spastic paresis, although some progress has recently been made on the search for a putative genetic component to the latter two syndromes.⁵⁰

Figure 3.8 Ultrasound‐guided collection of CSF from between C1 and C2 in a standing sedated horse (A and B). In C, the spinal cord (SC) and dura mater (green arrows) are imaged using a transversely oriented ultrasound transducer. A 3.5inch (8‐10 cm) 18 g styletted needle is introduced from below the transducer and advanced into the subarachnoid space, then the stylet is removed and CSF is aspirated into a syringe attached to extension tubing. The dashed orange line in C indicates a typical dorsolateral to ventromedial path of needle placement. The “Z” symbol at the top marks the dorsal aspect of the ultrasound image.

Source: Images courtesy of Sally DeNotta, University of Florida, USA.

With most CNS malformations, the CSF analysis will be normal, although those anomalies that result in tethering of neural tissues such as complex vertebral deformations can result in progressive disease with ongoing traction injury to CNS neural tissues during growth, thus potentially resulting in CSF changes consistent with trauma. If a malformation of the calvaria or vertebral column damages underlying nervous tissue, the CSF may reflect compression with evidence of subtle hemorrhage.

Infectious diseases can result in CSF pleocytosis and the elevation of protein content. The cell type present varies considerably, but generally neutrophils predominate with bacterial diseases and small mononuclear cells with viral diseases. Notable exceptions to this are high neutrophil numbers with Eastern equine encephalitis and high mononuclear cell numbers with listeriosis and equine neuroborreliosis. Fungal and protozoal diseases usually cause mixed cell responses. Protozoal, and particularly helminth parasite infestations, may produce an eosinophilic and neutrophilic response in the CSF, as well as hemorrhage. In contrast to bacterial meningitis where the neutrophils are degenerate and show toxic changes, with parasitic invasions and specific viral diseases the polymorphonuclear cells are nondegenerate and multilobulated due to their age. In most chronic inflammatory states and in diseases in which there is much CNS tissue necrosis, the CSF can contain many large mononuclear cells or macrophages.

Although immune‐associated CNS disorders such as canine steroid‐responsive meningitis have not yet been reported in large animals, there would be an expected modest pleocytosis, usually mononuclear, and, extrapolating from canine practice, it may well be worth sampling CSF from both the cervical and lumbar regions to maximize the likelihood of identifying the major CSF cytologic response.⁵¹ This recommendation also likely holds true for many of the infectious spinal disorders.

With traumatic injury to the CNS, there is often some hemorrhage into the CSF with resulting yellow discoloration that can persist for days after the insult. This xanthochromia remains after red cells have been centrifuged off. Neutrophils, not showing toxic changes, followed by macrophages, will usually appear in the CSF in response to hemorrhage.

In most toxic, nutritional, and metabolic neurologic diseases, the results of routine CSF analyses are generally normal. However, for those diseases in which there can be considerable tissue destruction, such as lead poisoning, sodium salt/water intoxication and polioencephalomalacia in ruminants, and moldy corn‐associated leukoencephalomalacia in horses, there may be some protein leakage into, and a mononuclear cell response within, the CSF.

Typically, there is leakage of protein and some xanthochromia without any significant pleocytosis in many vascular diseases. If the hemorrhage is large, then neutrophils and macrophages may also be seen.

Primary genetic (degenerative) diseases do not typically cause any changes in CSF constituents, and most frequently, the CSF analysis is normal. Early in the course of cell degeneration and especially in very young patients, some protein and even cellular response may be expected. Those resulting in the accumulation of products such as occurs in the storage diseases may result in macrophages containing waste material in the CNS and thence in CSF.

Neoplasms can act like other space‐occupying lesions, such as abscesses, granulomas (such as cholesterinic granulomas in horses) and hematomas, and can increase intracranial pressure. The most frequent CSF change in patients with neoplasia is a slight elevation in protein content. Sometimes there is evidence of mild injury, xanthochromia, and a few macrophages. Rarely, there have been atypical lymphocytes detected in CSF from cattle with CNS lymphosarcoma. Atypical cells such as melanoblasts have been detected in CSF samples, but it is worth considering whether such cells may have been disrupted from meningeal sites during the course of CSF collection.

Because of the lymphatic‐like drainage system of the CNS from perivascular and Virchow–Robin spaces ultimately to the subarachnoid space, any process that is contained within the parenchyma of the CNS may ultimately cause the leakage of pigments or protein, or the exfoliation of cells into the CSF.