Introduction

Cerebrospinal fluid (CSF) sampling, for measurement of cell count, cytological examination and total protein, can be thought of as the central nervous system (CNS) equivalent of serum biochemistry and haematological examination of peripheral blood. CSF analysis is extremely sensitive for the diagnosis of CNS disease but is rarely specific. It is a technique that can be performed easily in practice. Care needs to be taken to process the sample appropriately so that useful results are obtained.

When?

There are some key clinical situations in which CSF analysis is an extremely important diagnostic tool. Broadly speaking, these are mainly on suspicion of an encephalopathy or myelopathy; particularly if CNS inflammatory/infectious disease or CNS neoplasia is suspected. CSF abnormalities occasionally occur in other CNS disease processes but are rarely specific.

What clinical clues lead to a suspicion of CNS inflammation or infection? Animals that present with multifocal CNS signs, particularly when they include signs of meningeal pain, e.g., cervical pain, especially on head ventroflexion, are a key group. These animals may or may not be pyrexic, and may or may not have an inflammatory leucogram. It should be noted that it is quite common for animals with CNS inflammation to have no overt systemic evidence of inflammation.

CSF changes may also be seen when nerve roots are inflamed even if there is no direct CNS involvement, because the proximal nerve roots are encased by the meninges. The common clinical example of this is polyradiculoneuritis, which can be idiopathic or due to infectious agents such as Neospora or Toxoplasma.

Some of the most common CNS inflammatory/infectious diseases in the UK include: meningoencephalitis of unknown aetiology, including the syndrome known as granulomatous meningoencephalomyelitis (GME); steroid-responsive meningitis-arteritis (SRMA); meningoencephalitis secondary to feline infectious peritonitis (FIP); neosporosis; and toxoplasmosis.

Neoplastic changes commonly cause mild increases in CSF total protein and mildly raised cell counts. It is rare for CNS neoplastic lesions to exfoliate into CSF and so cytological findings are usually non-specific.

Contraindications for CSF collection are:

 Atlantoaxial subluxation or cervical vertebral fracture (for cisternal collection).

 If raised intracranial pressure (ICP) is suspected and CSF results are not required for a definitive diagnosis.

 If cerebellar herniation secondary to raised ICP has been observed on magnetic resonance imaging (MRI).

 If general anaesthesia is contraindicated.

How do we decide if ICP is raised in the clinical situation? It is uncommon to measure ICP in the clinic, and so a decision needs to be made on clinical grounds. First, the clinician needs to carefully consider the most likely differential diagnoses in a particular patient. If a large space-occupying lesion or hydrocephalus are high up on the list, CSF is best avoided unless advanced imaging can be done first. Secondly, the animal's neurological status needs to be assessed carefully. If an animal has a normal mental status, no pupillary abnormalities and good motor function, ICP is unlikely to be raised significantly. However, the safest method is to always perform MRI before CSF sampling. There is no evidence that lumbar puncture is any safer than cisternal sampling in animals with raised ICP.

How?

The cerebellomedullary cistern site (cisternal puncture) is the easiest way to achieve a successful diagnostic CSF tap. Lumbar collection is also possible but is more technically difficult and is more commonly associated with blood contamination or failure to collect sufficient volume. It is advisable to choose lumbar puncture if the lesion is suspected to be caudal to the cerebellomedullary cistern, because abnormalities are more likely to be found in CSF taken caudal to the lesion. 1 ml per 5 kg of body weight can be safely removed and should be collected into a sterile plain tube. The skin should be shaved and prepared aseptically.

Key Points Regarding Cisternal CSF Collection

 It is advisable to practise this technique on several cadavers before collecting CSF in a live animal.

 General anaesthesia is always required.

 The cuff of the endotracheal tube must be deflated or a non-collapsible tube should be used.

 The animal needs to be in right lateral recumbency for a right-handed operator with the head flexed at right angles and the nose held parallel to the table.

 Needle position: an imaginary line is drawn between the cranial limits of the wings of C1 and a perpendicular line from the external occipital protruberance caudally. The needle should be placed at the intersection of these lines.

 A change in texture, often described as a pop, is felt as the needle enters the subarachnoid space.

 The author prefers to use a 22 gauge or 20 gauge spinal needle, with stylet, of length 38 mm (1.5 inches) for most cisternal punctures. A 63 mm (2.5 inch) or 88 mm (3.5 inch) needle is sometimes required for cisternal puncture in giant breeds and most lumbar punctures.

 The author does not recommend aspirating the CSF as there is risk of damaging neural tissue. The CSF should be collected as it drips out of the needle hub.

Trouble-Shooting Cisternal CSF Collection

 The needle hits bone: reposition more cranially or more caudally.

 Pure blood emerges from the needle: the needle is probably off midline. Reposition.

Key Points Regarding Lumbar CSF Collection

 The animal needs to be in lateral recumbency with the pelvic limbs advanced cranially.

 The key to success with this technique is to palpate the dorsal spinous processes very carefully. In most dogs and cats, an L5/L6 puncture is advised, but L4/L5 is easier in large or obese dogs. The needle should be placed on the dorsal spine of L6 and walked cranially, while keeping precisely to midline, until the needle enters the space. The needle is then pushed through the cauda equina. A twitch of the legs or tail is commonly, but not always, encountered. Once the needle is against the ventral floor of the vertebral canal it needs to be withdrawn slightly and slowly until CSF emerges from the hub of the needle. Blood contamination or failure to collect sufficient volume are relatively common problems.

 The author does not recommend aspirating the CSF as there is risk of damaging neural tissue. The CSF should be collected as it drips out of the needle hub.

Trouble-Shooting Lumbar CSF Collection

 The needle repeatedly hits bone without entering vertebral canal: ask an assistant to flex the pelvic limbs more and check landmarks carefully. It is particularly important to remain in midline and start at the level of L6 (or L5) dorsal spinous process.

 Landmarks are only felt with great difficulty: take a radiograph to confirm position of needle and then reposition as necessary.

 A twitch is felt but CSF does not flow: withdraw the needle slightly.

Tests

Minimum Database

 Total and differential cell count

 Cytological examination

 Total protein concentration

Additional Tests If Indicated

 Polymerase chain reaction (PCR) for canine distemper virus (CDV), Neospora, FIP and Toxoplasma

 Bacterial culture

 Antibody titres, e.g., for canine distemper paired with serum, or IgA paired with serum

Why?

 To aid with definitive diagnosis

 To rule out specific diseases

Normal CSF Parameters

 Clear and colourless, with consistency of water.

 White blood cell count <5 WBC/µl in cisternal puncture, often less from lumbar puncture. The cells should be predominantly mononuclear.

 Total protein <0.25 g/l from cisternal puncture and approximately twice this value at the lumbar site.

Typical CSF Findings in Specific Disease Processes

 Markedly raised protein and neutrophilic pleocytosis in meningoencephalitis caused by FIP.

 Neutrophilic pleocytosis in steroid-responsive meningitis-arteritis.

 Mixed pleocytosis in granulomatous meningoencephalomyelitis and other miscellaneous meningoencephalitides.

 Mixed pleocytosis in neosporosis, toxoplasmosis, CDV.

 Marked degenerate neutrophilic pleocytosis in bacterial meningitis (rare) plus possible bacteria and positive culture.

 Mild pleocytosis and raised protein in: disc herniation, degenerative myelopathies, syringohydromyelia, following myelography.

 Xanthochromia (yellow tinge) following CNS haemorrhage.

None of these abnormalities is entirely specific for each disease, but they tend to be the most typical findings.

CSF Handling in Practice

It is very important that cytological examination is done within 30 minutes of collection. Sedimentation of CSF is a very useful method of preparing a slide for CSF cytology:

1.  Prepare a cylinder (1-2 cm diameter), e.g., from a blood tube or 5ml syringe, and attach to a microscope slide using paraffin wax.

2.  Place 1 ml of fresh CSF into tube and allow to sediment for 30 minutes.

3.  Remove supernatant and remove cylinder. Air-dry the slide and submit to the laboratory for examination.

References

1.  Ducote JM, Dewey CW. In: Dewey CW. ed. A Practical Guide to Canine and Feline Neurology. Iowa: Blackwell, 2003; 57-64.

2.  Wamsley H, Alleman AR. Clinical pathology. In: Platt, SR; Olby, NJ. eds. BSAVA Manual of Canine and Feline Neurology. Gloucester, UK: British Small Animal Association, 2004; 35-53.