Skip to main content
SpringerLink
Log in

Calcitonin gene-related peptide, substance P and GAP-43/B-50 immunoreactivity in the normal and arthrotic knee joint of the mouse

  • Published:
Histochemistry Aims and scope Submit manuscript

Abstract

The aim of this study was to describe the normal distribution of calcitonin gene-related peptide (CGRP) and substance P (SP) containing fibres in the knee joint of the mouse and to obtain insight into the changes in innervation associated with degenerative processes in the joint. Arthrosis was induced by a single subpatellar intra-articular injection of bacterial collagenase. After decalcification in EDTA solutions, the CGRP and SP fibres were visualized by peroxidase-antiperoxidase pre-embedding immunocytochemistry for light microscopy. Control experiments on the mouse brain as a reference for the effect of EDTA on the immunostaining showed that the decalcification procedure with EDTA had not impaired the immunostaining. A rich innervation of thin varicose CGRP and SP immunoreactive fibres was found in most peri- and intra-articular tissue components. The periosteum, synovial tissues, the joint capsule and the intra-articular fat tissues were richly innervated. Less intense innervations were also found in the subchondral bone plates of the tibio-femoral joint and of the patella. Fibres were also found in the soft tissues between the patellar tendon and the femoral groove. No differences could be found between the location of CGRP and SP fibres with respect to the localization in the joint, but generally more CGRP fibres were found. The collagenase-induced osteoarthrosis was characterized by sclerosis of the subchondral bone, patellar dislocation, osteophyte formation, synovial proliferation and by severe cartilage abrasion, particularly on the medial side of the femoro-tibial joint. The overall distribution of CGRP and SP fibres was the same as in the control joints. However, major differences were found in all studied joints at specific locations around the cruciate ligaments, in the synovium around the patella, in the soft tissues lateral of the patella and in plica tissue between the patella and femoral groove. The CGRP and SP innervation was no longer detectable by immunolabelling with the antibodies. With a polyclonal antibody to the growth associated protein GAP-43/B-50, signs of degenerated axonal profiles were observed in these locations. At other peripheral locations, such as the muscles, the GAP-43/B-50 distribution was normal. In conclusion, the present study provides detailed information on the localization of CGRP and SP fibres, which may be involved in pain perception. Knowledge of the changes that occur during arthrosis may give more insight into the clinical symptoms.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  • Arnoldi CC, Lemperg R, Linderholm H (1975) Intraosseous hypertension and pain in the knee. J Bone Joint Surg [Br] 57:360–363

    Google Scholar 

  • Bendele AM (1987) Progressive chronic osteoarthritis in femorotibial joints of partial medial menisectomized guinea pigs. Vet Pathol 24:444–448

    Google Scholar 

  • Bendele AM, Hulman JF (1988) Spontaneous cartilage degeneration in guinea pigs. Arthritis Rheum 31:561–565

    Google Scholar 

  • Benske J, Schünke M, Tillmann B (1988) Subchondral bone formation in arthrosis. Polychrome labeling studies in mice. Acta Orthop Scand 59:536–541

    Google Scholar 

  • Bentley G (1971) Papain-induced degenerative arthritis of the hips in rabbits. J Bone Joint Surg 53[Br]:324–337

    Google Scholar 

  • Bjurholm A, Kreicbergs A, Brodin E, Schulzberg M (1988a) Substance P-and CGRP-immunoreactive nerves in bone. Peptides 9:165–171

    Google Scholar 

  • Bjurholm A, Kreicbergs A, Terenius L, Goldstein M, Schultzberg M (1988b) Neuropeptide Y-tyrosine hydroxylase- and vasoactive intestinal polypeptide-immunoreactive nerves in bone and surrounding tissues. J Auton Nerv Syst 25:119–125

    Google Scholar 

  • Bjurholm A, Kreicbergs A, Schulzberg M (1989) Fixation and demineralization of bone tissue for immunohistochemical staining of neuropeptides. Calcif Tissue Int 45:227–231

    Google Scholar 

  • Brodin E, Gaselius B, Panapoulos P, Olgart I (1983) Morphine inhibits substance-P release from peripheral sensory nerve endings. Acta Physiol Scand 117:567–570

    Google Scholar 

  • Burgess PR, Clark FJ (1969) Characteristics of knee joint receptors in the cat. J Physiol 203:317–335

    Google Scholar 

  • Cavanaugh JM, El-Bohy A, Hardy WN, Getchell TV, Getchell ML, King AI (1989) Sensory innervation of soft tissues of the lumbar spine in the rat. J Orthop Res 7:378–388

    Google Scholar 

  • Ceder LC, Larson RL (1979) Z-plasty lateral retinacular release for the treatment of patellar compression syndrome. Clin Orthop 144:110–113

    Google Scholar 

  • Cervero F, Schaible H-G, Schmidt RF (1991) Tonic descending inhibition of spinal cord neurones driven by joint afferents in normal cats and in cats with an inflammed knee joint. Exp Brain Res 83:675–678

    Google Scholar 

  • Cooke TDV (1985) Pathogenetic mechanisms in polyarticular osteoarthritis. Clin Rheum Dis 11:203–238

    Google Scholar 

  • Cooke TDV, Chir B (1986) Immune pathology in polyarticular osteoarthritis. Clin Orthop Rel Res 213:41–49

    Google Scholar 

  • Farfan HF (1983) On the nature of arthritis. J Rheum 10 [Suppl 9]:103–104

    Google Scholar 

  • Ficat RP, Hungerford DS (1977) Disorders of the patellofemoral joint. Williams and Wilkins, Baltimore

    Google Scholar 

  • Ficat RP, Ficat C, Gedeon P, Toussaint JB (1979) Spongialization: a new treatment for deceased patellae. Clin Orthop 144:74–83

    Google Scholar 

  • Fulkerson JP, Schutzer SF (1986) After failure of conservative treatment for painful patellofemoral malalignment: lateral release or realignment? Orthop Clin North Am 17:283–288

    Google Scholar 

  • Fulkerson JP, Tennant R, Jaivin JS, Grunnet M (1985) Histologic evidence of retinacular nerve injury associated with patellofemoral malalignment. Clin Orthop 197:196–205

    Google Scholar 

  • Gardner E (1948) The innervation of the knee joint. Anat Rec 101:109

    Google Scholar 

  • Goldner J (1938) A modification of the Masson trichrome technique for routine laboratory purposes. Am J Pathol 14:247–249

    Google Scholar 

  • Goodfellow J, Hungerford DS, Woods C (1976) Patello-femoral joint mechanics and pathology. J Bone Joint Surg [Br] 58:291–299

    Google Scholar 

  • Gorgels TGMF, Oestreicher AB, Kort EJM de, Gispen WH (1987) Immunocytochemical distribution of the protein kinase C substrate B-50 (GAP 43) in developing rat pyramidal tract. Neurosci Lett 83:59–64

    Google Scholar 

  • Gorgels TGMF, van Lookeren Campagne M, Oestreicher AB, Gispen WH (1989) B-50/GAP43 is localized at the cytoplasmic side of the plasma membrane in developing and adult rat pyramidal tract. J Neurosci 9:3861–3869

    Google Scholar 

  • Grönblad M, Liesi P, Korkalo O, Karaharju E, Polak J (1984) Innervation of human bone periosteum by peptidergic nerves. Anat Rec 209:297–299

    Google Scholar 

  • Grönblad M, Korkala O, Liesi P, Karaharju E (1985) Innervation of synovial membrane and meniscus. Acta Orthop Scand 56:484–486

    Google Scholar 

  • Grönblad M, Konttinen YT, Korkala O, Liesi P, Hukkanen M (1988) Neuropeptides in synovium of patients with rheumatoid arthritis and osteoarthritis. J Rheumatol 15:1807–1810

    Google Scholar 

  • Gulbenkian S, Merighi A, Wharton J, Varndell IM, Polak JM (1986) Ultrastructural evidence for the coexistence of calcitonin gene-related peptide and substance P in secretory vesicles of peripheral nerves in the guinea pig. J Neurocytol 15:535–542

    Google Scholar 

  • Halata Z, Haus J (1989) The ultrastructure of sensory nerve endings in human anterior cruciate ligaments. Anat Embryol 179:415–421

    Google Scholar 

  • Hanesch U, Heppelmann B, Schmidt RF (1991) Substance P-and calcitonin gene-related peptide immunoreactivity in primary afferent neurons of the cat's knee joint. Neuroscience 45:185–193

    Google Scholar 

  • Haus J, Halata Z (1990) Innervation of the anterior cruciate ligaments. Int Orthop 14:293–296

    Google Scholar 

  • Henry JL (1976) Effects of substance-P on functionally identified units in cat spinal cord. Brain Res 114:439–451

    Google Scholar 

  • Henry JH, Craven PR (1981) Surgical treatment of patellar instability: indications and results. Am J Sports Med 9:82–85

    Google Scholar 

  • Heppelmann B, Schaible H-G (1990) Origin of sympathetic innervation of the knee joint in the cat: a retrograde tracing study with horseradish peroxidase. Neurosci Lett 108:71–75

    Google Scholar 

  • Hill EL, Elde R (1988a) VIP-containing nerve fibers in bone and periosteum may be adrenergic. Proc NY Acad Sci 527:643–645

    Google Scholar 

  • Hill EL, Elde R (1988b) Calcitonin gene-related peptide-immunoreactive nerve fibres in mandibular periosteum: evidence for primary afferent origin. Neurosci Lett 85:172–178

    Google Scholar 

  • Hill EL, Elde R (1991) Distribution of CGRP-, VIP-, DβH-, SP-, and NPY immunoreactive nerves, in the periosteum of the rat. Cell Tissue Res 264:469–480

    Google Scholar 

  • Hohmann EL, Elde RP, Rysavy JA, Einzig S en Gebhard RL (1986) Innervation of periosteum and bone by sympathetic vasoactive intestinal peptide-containing nerve fibers. Science 232:868–871

    Google Scholar 

  • Holzer P (1988) Local effector functions of capsaicin-sensitive sensory nerve endings: involvement of tachikins, calcitonin generelated peptide and other neuropeptides. Neuroscience 24:739–768

    Google Scholar 

  • Hope PJ, Jarrott B, Schaible H-G, Clarke RW, Duggan AW (1990) Release and spread of immunoreactive neurokinin A in the cat spinal cord in a model of acute arthritis. Brain Res 533:292–299

    Google Scholar 

  • Hukkanen M, Grönblad M, Rees R, Konttinen Y, Gibson SJ, Hietanen J, Polak JM, Brewerton DA (1991) Regional distribution of mast cells and peptide containing nerves in normal and adjuvant arthritic rat synovium. J Rheumatol 18:177–183

    Google Scholar 

  • Kar S, Gibson SJ, Ress RG, Jura WGZO, Brewerton DA, Polak JM (1991) Increased calcitonin gene-related peptide (CGRP), substance P, and enkephalin immunoreactivities in dorsal spinal cord and loss of CGRP-immunoreactive motoneurons in arthritic rats depend on intact peripheral nerve supply. J Mol Neurosci 3:7–18

    Google Scholar 

  • Kennedy JC, Alexander IJ, Hayes KC (1982) Nerve supply of the human knee and its functional importance. Am J Sports Med 10:329–335

    Google Scholar 

  • Kristjansson GS, Zwiers H, Oestreicher AB, Gispen WH (1982) Evidence that the synaptic phosphoprotein B-50 is localized exclusively in nerve tissue. J Neurochem 39:371–378

    Google Scholar 

  • Kruger L, Silverman JD, Mantyh PW, Sternini C, Brecha NC (1989) Peripheral patterns of calcitonin gene-related peptide and general somatic sensory innervation: cutaneous and deep terminations. J Comp Neurol 280:291–302

    Google Scholar 

  • Kuntz A, Richings CA (1945) Innervation of the bone marrow. J Comp Neurol 83:213–222

    Google Scholar 

  • Kurashi Y, Hirota N, Sugimoto M, Satoh M, Takagi H (1983) Effect of morphine: a noxious stimuli-induced release of substance-P from rabbit dorsal horn in vivo. Life Sci 33:693–696

    Google Scholar 

  • Linder JE (1978) A simple and reliable method for silver impregnation of nerves in paraffin sections of soft and mineralized tissues. J Anat 127:543–551

    Google Scholar 

  • Marshall KW, Tatton WG (1990) Joint receptors modulate short and long latency muscle responses in the awake cat. Exp Brain Res 83:137–150

    Google Scholar 

  • Milgran JW, Robinson RA (1965) An electronic microscopic demonstration of unmyelinated nerves in haversian canals of the adult dog. Bull Johns Hopkins Hosp 117:163–173

    Google Scholar 

  • Miller MR, Kasahara M (1963) Observations on the innervation of human long bones. Anat Rec 145:13–23

    Google Scholar 

  • Morscher F (1978) Osteotomy of the patella in chondromalacia. Arch Orthop Trauma Surg 92:139–147

    Google Scholar 

  • Nieuwenhuys R (1985) Chemoarchitecture of the brain. Springer, Heidelberg New York

    Google Scholar 

  • Oestreicher AB, Gispen WH (1986) Comparison of immunocytochemical distribution of the phosphoprotein B-50 in the cerebellum and hippocampus of immature and adult rat brain. Brain Res 375:267–479

    Google Scholar 

  • Otsuka M, Yanagisawa M (1987) Does substance P act as a pain transmitter? Trends Pharmacol Sci 8:506–510

    Google Scholar 

  • Outerbridge RF (1964) Further studies on the etiology of chondromalacia patellae. J Bone Joint Surg [Br] 46:179–190

    Google Scholar 

  • Reimann I, Christensen SB (1977) A histological demonstration of nerves in subchondral bone. Acta Orthop Scand 48:345–352

    Google Scholar 

  • Salo PT, Tatton WG (1991) Age-related loss of joint innervation. Proceedings of the combined meeting of the orthopaedic research society of USA, Japan, Canada

  • Schaible H-G, Jarrot B, Hope PJ, Duggan AW (1990) Release of immunoreactive substance P in the spinal cord during development of acute arthritis in the knee joint of the cat: a study with antibody microprobes. Brain Res 529:214–223

    Google Scholar 

  • Schünke M, Tillmann B, Brück M, Müller-Ruchholz W (1988) Morphologic characteristics of developing osteoarthrotic leasions in the knee cartilage of STR/IN mice. Arthritis Rheum 31:898–905

    Google Scholar 

  • Schutte MJ, Dabezies EJ, Zimny ML, Happel LT (1987) Neural anatomy of the human anterior cruciate ligament. J Bone Joint Surg 69[Am]:243–247

    Google Scholar 

  • Sherman MS (1963) The nerves of bone. J Bone Joint Surg 45[Am]:522–528

    Google Scholar 

  • Silverman JD, Kruger L (1987) An interpretation of dental innervation based upon the pattern of calcitonin gene-related peptide (CGRP)-immunoreactive thin sensory axons. Somatosensory Res 5:157–175

    Google Scholar 

  • Sjölander P, Johansson H, Sojka P, Rehnholm A (1989) Sensory nerve endings in the cat cruciate ligaments: a morphological investigation. Neurosci Lett 102:33–38

    Google Scholar 

  • Skene JHP (1989) Axonal growth-associated proteins. Annu Rev Neurosci 12:127–156

    Google Scholar 

  • Sojka P, Johansson H, Sjölander P, Lorentzon R, Djupsjöbacka M (1989) Fusimotor neurons can be reflexly influenced by activity in receptor afferents from posterior cruciate ligaments. Brain Res 483:177–183

    Google Scholar 

  • Sokoloff L (1956) Natural history of degenerative joints disease in small laboratory animals. I. Pathologic anatomy of degenerative joint disease in the mice. Arch Pathol 62:118–128

    Google Scholar 

  • Sokoloff L, Jay GE (1956) Natural history of degenerative joints disease in small laboratory animals. II. Epiphyseal maturation osteoarthritis of the knee of mice of inbred strains. Arch Pathol 62:129–135

    Google Scholar 

  • Sternberger LA (1979) Immunocytochemistry, 2nd edn. Wiley, New York

    Google Scholar 

  • Tetzlaff W, Zwiers H, Lederis K, Cassar L, Bisby MA (1989) Axonal transport and localization of B-50/GAP 43-like immunoreactivity in regenerating sciatic and facial nerves of the rat. J Neurosci 9:1303–1313

    Google Scholar 

  • Thurston TJ (1982) Distribution of nerves in long bones as shown by silver impregnation. J Anat 134:719–728

    Google Scholar 

  • Van der Kraan PM, Vitters EL, Van Beuningen HM, Van der Putte LBA, Van den Berg WB (1990) Degenerative joint lesions in mice after a single intra-articular collagenase injection. A new model of osteoarthritis. J Exp Pathol 71:19–31

    Google Scholar 

  • Van Lookeren Campagne M, Oestreicher AB, Buma P, Verkley AJ, Gispen WH (1991) Ultrastructural localization of adrenocorticotrophic hormone and the phosphoprotein B-50/growth-associated protein 43 in freeze-substituted, lowicryl HM20-embedded mesencephalic central gray substance of the rat. Neuroscience 42:517–529

    Google Scholar 

  • Venesio T, Mulatero B, Fasolo A (1987) Coexistence of substance P and calcitonin gene-related peptide in the frog spinal cord. Neurosci Lett 80:246–250

    Google Scholar 

  • Verhaagen J, Oestreicher AB, Edwards PM, Veldman H, Jennekens FGI, Gispen WH (1988) Light- and electron-microscopical study of phosphoprotein B-50 following denervation and reinnervation of the rat soleus muscle. J Neurosci 8:1759–1766

    Google Scholar 

  • Villanueva AR (1988) A new Goldner's one step trichrome stain for identification of osteoid seams, bone and cells in undecalcified plastic embedded sections of bone. J Histotechnol 11:249–251

    Google Scholar 

  • Wakisaka S, Ichikawa H, Nishimoto T, Matsuo S, Yamamoto K, Nakata T, Akai M (1986) A technique for decalcification and demonstration of substance P-like immunoreactivity. Stain Technol 61:39–42

    Google Scholar 

  • Walton M (1977a) Degenerative joint disease in the mouse knee: radiological and morphological observations. J Pathol 123:97–107

    Google Scholar 

  • Walton M (1977b) Degenerative joint disease in the mouse knee: histologic observations. J Pathol 123:109–122

    Google Scholar 

  • White D, Helme R (1985) Release of substance-P from peripheral nerve terminals following electrical stimulation of the sciatic nerve. Brain Res 336:27–31

    Google Scholar 

  • Widenfalk G, Wiberg M (1989) Origin of sympathetic and sensory innervation of the knee joint. A retrograde axonal tracing study in the rat. Anat Embryol 180:317–323

    Google Scholar 

  • Williams JM, Brandt KD (1984) Immobilization ameloriates chemically-induced articular cartilage damage. Arthritis Rheum 27:208–216

    Google Scholar 

  • Wojtys EM, Beaman DN, Glover RA, Janda D (1990) Innervation of the human knee joint by substance-P fibers. J Arthroscopy Rel Surg 6:254–263

    Google Scholar 

  • Zimny ML, Schutte M, Dabezies E (1986) Mechanoreceptors in the human anterior cruciate ligament. Anat Rec 214:204–209

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Orthopaedics, Laboratory for Experimental Orthopaedics, Section Histomorphology, University Hospital Nijmegen, P.O. Box 9101, 6500 HB, Nijmegen, The Netherlands

    P. Buma, C. Verschuren & D. Versleyen

  2. Department of Rheumatology, Laboratory for Experimental Rheumatology, University Hospital Nijmegen, P.O. Box 9101, 6500 HB, Nijmegen, The Netherlands

    P. Van der Kraan

  3. Division of Molecular Neurobiology, Rudolf Magnus Institute and Institute of Molecular Biology and Medical Biotechnology, University of Utrecht, 3584 CH, Utrecht, The Netherlands

    A. B. Oestreicher

Authors
  1. P. Buma
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. C. Verschuren
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. D. Versleyen
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. P. Van der Kraan
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. A. B. Oestreicher
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Buma, P., Verschuren, C., Versleyen, D. et al. Calcitonin gene-related peptide, substance P and GAP-43/B-50 immunoreactivity in the normal and arthrotic knee joint of the mouse. Histochemistry 98, 327–339 (1992). https://doi.org/10.1007/BF00270017

Download citation

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00270017

Keywords

Search

Navigation