A silver-impregnation and immunocytochemical study of innervation of the distal sesamoid bone and its suspensory ligaments in the horse.

The innervation of the navicular bone (os sesamoideum distale) and its suspensory ligaments (ligamenta sesamoidea collateralia) (CSL) or proximal suspensory ligament and the ligamentum sesamoideum distale impar or the distal sesamoidean impar ligament (DS-impar ligament) was examined using combined anatomical techniques of silver impregnation and immunocytochemistry. Silver impregnation studies revealed an abundance of nerve fibres present in both the CSL and DS-impar ligament with the latter having relatively more nerve fibres. These silver-impregnated nerves coursed parallel to and were associated with the vasculature rather than appearing to innervate the vessels. Immunocytochemistry identified several sensory-related neuropeptides (calcitonin gene-related peptide (CGRP), substance P (SP) and neurokinin A (NKA)) in the nerves of the navicular bone and suspensory ligaments. More peptidergic nerves were evident within the synovial membrane and loose connective tissue in the dorsal part than in the palmar aspect of the CSL. In the CSL along the synovial membrane bordering the distal interphalangeal joint, the CGRP, SP and NKA were present in the nerves of vessels as well as the intimal layer of the distal interphalangeal joint. In the DS-impar ligament, there were many more nerves innervating vessels and the synovial membrane between the navicular bone and the third phalanx than were present in these structures in the CSL. Nerves with all 3 peptides entered the navicular bone via the proximal border and the distal groove to innervate the perichondrium, trabeculae and osteons. SP-like nerves also innervated the cortical bone underlying the articular cartilage. We suggest that these sensory nerve peptides contribute to the pathology of the navicular syndrome. The distribution of the nerves in the CSL and the DS-impar ligament could explain the clinical effects of local anaesthetics injected into the distal interphalangeal joint.

Immunocytochemical and dye distribution studies of nerves potentially desensitized by injections into the distal interphalangeal joint or the navicular bursa of horses.

To determine whether the distal interphalangeal (DIP) joint directly or indirectly communicates with the navicular bursa (bursa podotrochlearis) and to identify sensory nerves in these synovial structures that might be desensitized by intra-articular injections of anesthetics, Evans blue dye in physiologic saline solution, Luxol fast blue dye with mepivicaine, or commercial latex was injected into the DIP joint (5 ml) or the navicular bursa (3 ml) of 152 digits obtained from horses or ponies at necropsy. The digits were frozen, cut with a band saw, and examined for distribution of dye or latex. Of 122 digits that had injections into the DIP joint, 120 did not have evidence of a communication between the DIP joint and either the navicular bursa or digital flexor tendon sheath. Of 16 digits that had injections into the navicular bursa, 14 did not have evidence of a direct communication with the DIP joint. Injection of dye into the DIP joint resulted in diffusion of dye and staining of other structures, including the synovial linings of the collateral sesamoidean ligaments and of the distal sesamoidean impar ligament and the medullary cavity of the navicular bone. In addition, a blue tinge was observed in the navicular bursa after dye was injected into the DIP joint, suggesting an indirect, and potentially functional, communication between the DIP joint and the navicular bursa. Injection of dye into the navicular bursa resulted in staining only of the bursa's synovial lining. Immunocytochemical analysis revealed nerves immunoreactive for the peptidergic neurotransmitters substance P, and calcitonin gene-related peptide located in structures that were stained after dye was injected into the DIP joint.(ABSTRACT TRUNCATED AT 250 WORDS)

Substance P innervation of equine synovial membranes: joint differences and neural and nonneural receptor localizations.

Substance P (SP) immunocytochemistry and receptor autoradiography were used to define the innervation of the equine synovial membrane of joints equivalent to the wrist and knuckle of man. SP-immunoreactive fibers were mainly concentrated around blood vessels in the subsynovial layer, although not exclusively, while in the more distal joint, SP fibers were more frequently seen in the synovial surface layer. Iodinated SP receptor autoradiography studies revealed silver grain concentrations in the advential layer of blood vessels associated with the vasa vasorum, on the vascular endothelium and in the synovial surface. These findings suggest that SP has various sites of action within the synovial membrane, each of which may contribute both a sensory function and a different component of the inflammatory process to the joint.

Sensory receptors in the equine foot.

Two types of sensory receptors were located in the equine foot, using anatomic techniques. Histologic examination of stained hoof sections revealed lamellated corpuscles in the hoof dermis, which had many of the morphologic characteristics of Pacinian corpuscles. These sensory receptors were restricted to the palmar (caudal) aspects of the solar dermis of the heel. A second type of receptor was detected by use of immunocytochemistry, indicating apparently naked nerve endings containing the neuropeptide calcitonin gene-related peptide-like immunoreactivity in skin, solar dermal tubules, and the digital cushion. This peptide is an example of a sensory neurotransmitter contained in dorsal root ganglion cells and is believed to exist only in unmyelinated sensory nerve fibers. These 2 morphologic structures may be used for detection of sensory stimuli, such as pressure (or vibratory senses) and pain, respectively, in horses during various locomotory gaits.

Neuropeptidergic innervation of equine synovial joints.

Immunocytochemical analysis of equine synovial membranes revealed presence of several neuropeptides, including substance P (SP), neurokinin A, and neuropeptide Y, in nerves of the radiocarpal, middle carpal, and metacarpophalangeal (fetlock) joints. Within the subsynovium, these neuropeptides were located perivascularly, whereas in the fronds, only neuropeptide Y was restricted to the vessels of the synovial membrane. Only SP and neurokinin A were found in the intimal layer. The intimal layer of the metacarpophalangeal joint contained more SP-immunoreactive fibers than were observed in the intimal layer of the radiocarpal joint. Substance P also was detected in the synovial fluid from all 3 joints, but mean +/- SD concentrations were significantly different only between the middle carpal joint (37.56 +/- 5.48 fmol/ml; n = 6) and the metacarpophalangeal joint (55.80 +/- 8.33 fmol/ml; n = 5) and between the middle carpal joint and the radiocarpal joint (52.43 +/- 14.60 fmol/ml; n = 7).

Substance P in the synovial membrane and fluid of the equine middle carpal joint.

This preliminary study was designed to determine whether the neurotransmitter substance P was present in the middle carpal synovial membrane of the normal horse and whether the neuropeptide could be identified in the synovial fluid of normal horses and those with joint diseases. Immunocytochemistry on middle carpal synovial membrane biopsies from fresh cadavers was used to demonstrate substance P-containing neural elements. Substance P was most abundant in the subintimal portion of the membrane, with occasional filaments coursing via synovial fronds to the intimal portion. Radioimmunoassay techniques were used on acidified acetonitrile-preserved synovial fluid samples to measure substance P concentrations. Fluid from 9 joints of 5 normal horses and 6 joints of 4 horses with joint diseases were analysed. Disease conditions included acute and chronic osteoarthritis and osteochondrosis. Synovia from normal horses contained a mean concentration of substance P significantly less than that of horses with joint diseases (P less than 0.05). Elevated concentrations of neurotransmitters in diseased joints suggests a potential contribution to the pathophysiology of joint disorders in horses.