The Pain State Arising From the Laminitic Horse: Insights Into Future Analgesic Therapies

Inflammation 

On the basis of clinical signs and the chemical milieu, it has been supposed that the laminitic hoof represents an inflammatory model. The laminitic hoof displays the presence of products arising from blood (complement) and blood products (lymphocytes, platelets), circulating and local inflammatory cells (neutrophils and macrophages), and endothelial cells. Given the presence of these cellular elements, it is not surprising that agents, such as complement and kinins (from plasma), serotonin from platelets, cytokines (tumor necrosis factor [TNF] and interleukin-1ß) and cyclooxygenase products (prostaglandins, leukotrienes) from inflammatory cells, endothelin (vasculature), and free radicals (neutrophils) have been variously identified in the laminitic hoof.4, 5, 6

An important property of the high-threshold afferent C fibers is that they are imminently responsive to virtually all the chemical products outlined earlier in the text.7 In a variety of organ systems, these products can act though eponymous receptors located on the terminal of high-threshold C fibers and initiate two events: (i) the first is to produce an activation of the C-fiber terminal, with the frequency of activation proportional to local concentrations of the many hormones and (ii) through intraterminal transduction cascades to sensitize the terminal such that its threshold for activation by mechanical stimuli (e.g., pressure) is substantially reduced. Accordingly, relatively low-intensity stimuli become adequate to activate a nociceptive sensory axon.8 These events lead to ongoing afferent traffic to the spinal cord and a shift to the left in the stimulus-frequency response curve, indicating that for any given stimulus, the axon shows a greater response.

In addition, to this peripheral sensitization, the ongoing small afferent input initiates a central state of facilitation mediated in part by the spinal release of excitatory amino acids (glutamate), tachykinins (substance P), cyclooxygenase products (prostaglandin E2), chemokines (fractalkine), and cytokines (TNF, interleukin-1ß). Some of these products derive from sensory afferents as well as local neurons and non-neuronal cells (astrocytes and microglia). These products released by persistent afferent input will act upon eponymous receptors and lead to spinal synaptic excitability. These dorsal horn cascades have been reviewed in detail elsewhere.9, 10, 11, 12 It is currently believed that it is this joint peripheral and spinal facilitation which underlies an inflammation evoked hyperalgesic state.

Nerve Injury 

While it is evident that inflammation and peripheral sensitization are relevant to the mechanisms of pain in the laminitic hoof, more recent evidence has suggested the probable contributions of nerve injury. Recent work by Jones et al13 has shown in the lateral digital nerves that laminitis is associated with both significant morphological abnormalities and losses of both myelinated and unmyelinated axons. In the C8 DRG (which innervates the front hoof) there was an increased expression of Activation Transcription Factor (ATF3), a marker of peripheral nerve injury in both large and small afferents. The specificity of these changes is suggested by the fact that these changes in Activation Transcription Factor (ATF3) expression were not observed in the C4 ganglion (which does not innervate the hoof). In addition, there was an increased expression of Neuropeptide Y in these DRG cells. As reviewed elsewhere,14 these somatotopically linked changes in nerve and DRG have been commonly observed in preclinical models of pain secondary to peripheral nerve injury. In these models, changes in axon morphology and DRG expression have been shown to signal long-term changes in peripheral and central processing. These changes include: (i) Increased Persistent (ectopic) afferent activity arising from the injured terminal and DRG of the injured nerve15; (ii) The ectopic activity represents the local increase in the expression of sodium channels in the DRG and axons of afferent innervating the laminitic hoof and increasing sensitivity to excitatory products, such as cytokines released from local inflammatory cells8; (iii) Loss of dorsal horn inhibition otherwise mediated by dorsal horn gamma amino butyric acid (GABA) and glycine chloride ionophores15; (iv) Sympathetic sprouting leading to an increased excitatory coupling with the peripheral neuroma and the associated dorsal root ganglia by postganglionic sympathetic terminals16; and (v) Activation of astrocytes and microglia leading to an increased spinal expression of proexcitatory products.17, 18

While there has been little direct assessment of the pain biology associated with the laminitic hoof, the limited picture outlined above is consistent with a complex pain state involving both an inflammatory and perhaps less expectedly, a neuropathic component. This picture in animal models would be consistent with the sensitization of high-threshold polymodal nociceptors, the development of ectopic activity in from nerves innervating the hoof and from the dorsal root ganglia. In addition, biological markers to be expected in the DRG and spinal cord segments receiving afferent input form the laminitic hoof. These changes would include increased expression of subtypes of voltage sensitive sodium channels, increased expression of subunits of voltage sensitive calcium channels (e.g., α2δ), the binding site for the anticonvulsant gabapentin,19 of the laminitic animals would be changes in the excitatory receptors origin dorsal horn pain processing, notably those for the NMDA and AMPA glutamate receptors, and marked appearance of markers for astrocyte (glial fibrillary acidic protein) and microglial (OX42) activation. Future histological studies are required to address those possibilities. An important question clearly relates to the time course of the evolving disease, and the possibility that at certain early stages, the phenomena may be largely associated with nociceptor sensitization; while in a chronic condition, the inflammatory environment leads to changes which initiate the neuropathic elements observed by Jones et al. (2007). Thus, would there be a difference between the laminitic acute state observed after precipitating exposure to black walnut or over feeding versus a persistent state secondary to altered patterns of weight bearing?

Endothelin 

This is a peptide that is present in endothelial, mast cell, monocytes macrophages keratinocytes, and neutrophils. It is released in the face of local inflammation, and is increased in laminitis and acts through several endothelin receptors. These receptor locations are present in small DRG cell (small, primary afferents), immune cells, keratinocytes, and endothelial cells. In the face of peripheral inflammation, there is an increased expression. This agent is known to produce hyperpathia, and inhibitors (ETA) have been shown to reduce hyperpathia.22

Tumor Necrosis Factor 

TNF is located on monocytes macrophages, and neutrophils. Acting through one of several TNF receptors (primarily TNF receptor-1), it has been shown to activate terminals and nerves. In the face of inflammation and nerve injury, there is an increase in its expression in the injury site and in the DRG of the injured axon. Activation of TNF receptor-1 yields hyperpathia. Antisense (TNF) and decoy protein has been shown to reduce hyperpathia in a variety of inflammatory and neuropathic pain states.22

Sodium Channels 

After nerve injury, there is an increase in the expression of subtypes of voltage sensitive sodium channels, and these are responsible for ectopic activity in the injured nerve. Systemic (subcutaneous/intravenous) lidocaine given at doses which produce plasma levels of 1–3μg/mL, concentrations below those producing cardiac effects or blocking normal axon conduction, can reduce that ectopic activity and lead to a reversal of the neuropathic pain state.23 Afferent axon conduction is dependent on voltage-gated sodium channels. Future direction are focused on agents that block tetrodotoxin-insensitive channels that are found largely on C fibers. None are as yet commercially available.24 An alternative is the possible development of techniques to permit specific block of axons bearing the TRPV1 receptor, to permit entry of local anesthetics that cannot otherwise penetrate axon membranes.25 These approaches would permit, in theory, perineural block of a mixed nerve limited to C fibers.

Primary Afferent TRPV1 Receptor 

High-threshold peptidergic C fibers express TRPV1 (+) receptors. They are believed to be a critical link in nociceptive traffic. TRPV-1 agonists such as capsaicin or resniferotoxin activate and then desensitize the terminal (peripheral or spinal) and/or deplete its transmitter and then block C fiber transmission. Peripheral perineural application of capsaicin has been shown to block a variety of pain states.26, 27 Intrathecal delivery of resniferotoxin in dogs has been shown to produce a persistent and clinically relevant pain relief in dogs with osteoarthritis and osteosarcoma.28

Neurokinin 1 Receptor Toxins 

Neurokinin 1 receptors are located on dorsal horn projection neurons known to be essential for nociception. Substance P coupled to the 31kDA saporin will bind to the neurokinin 1 receptor, become internalized, and kill the cell, as it blocks ribosylation. Preclinical studies in rats and dogs has shown that it produces a persistent antinociception in the face of inflammatory and neuropathic pain. The toxin is selective as it will only destroy cells that bear the Nk1 receptor.29 Intrathecal delivery of sP-saporin in dogs has been shown to produce a persistent loss of lamina pain projection neurons in the canine model.30