Effects on hemodynamic enhancement and discomfort of a new textile electrode integrated in a sock during calf neuromuscular electrical stimulation.

PURPOSE: To compare fixed transverse textile electrodes (TTE) knitted into a sock versus motor point placed standard gel electrodes (MPE) on peak venous velocity (PVV) and discomfort, during calf neuromuscular electrical stimulation (calf-NMES). METHODS: Ten healthy participants received calf-NMES with increasing intensity until plantar flexion (measurement level I = ML I), and an additional mean 4 mA intensity (ML II), utilizing TTE and MPE. PVV was measured with Doppler ultrasound in the popliteal and femoral veins at baseline, ML I and II. Discomfort was assessed with a numerical rating scale (NRS, 0-10). Significance was set to p < 0.05. RESULTS: TTE and MPE both induced significant increases in PVV from baseline to ML I and significantly higher increases to ML II, in both the popliteal and femoral veins (all p < 0.001). The popliteal increases of PVV from baseline to both ML I and II were significantly higher with TTE versus MPE (p < 0.05). The femoral increases of PVV from baseline to both ML I and II were not significantly different between TTE and MPE. TTE versus MPE resulted at ML I in higher mA and NRS (p < 0.001), and at ML II in higher mA (p = 0.005) while NRS was not significantly different. CONCLUSION: TTE integrated in a sock produces intensity-dependent increases of popliteal and femoral hemodynamics comparable to MPE, but results in more discomfort at plantar flexion due to higher current required. TTE exhibits in the popliteal vein higher increases of PVV compared to MPE. TRIAL REGISTRATION: Trial_ID: ISRCTN49260430. Date: 11/01/2022. Retrospectively registered.

Neuromuscular electrical stimulation in garments optimized for compliance.

PURPOSE: Physical inactivity is associated with muscle atrophy and venous thromboembolism, which may be prevented by neuromuscular electrical stimulation (NMES). This study aimed to investigate the effect on discomfort, current amplitude and energy consumption when varying the frequency and phase duration of low-intensity NMES (LI-NMES) via a sock with knitting-integrated transverse textile electrodes (TTE). METHODS: On eleven healthy participants (four females), calf-NMES via a TTE sock was applied with increasing intensity (mA) until ankle-plantar flexion at which point outcomes were compared when testing frequencies 1, 3, 10 and 36 Hz and phase durations 75, 150, 200, 300 and 400 µs. Discomfort was assessed with a numerical rating scale (NRS, 0-10) and energy consumption was calculated and expressed in milli-Joule (mJ). Significance set to p ≤ 0.05. RESULTS: 1 Hz yielded a median (inter-quartile range) NRS of 2.4 (1.0-3.4), significantly lower than both 3 Hz with NRS 2.8 (1.8-4.2), and 10 Hz with NRS 3.4 (1.4-5.4) (both p ≤ .014). Each increase in tested frequency resulted in significantly higher energy consumption, e.g. 0.6 mJ (0.5-0.8) for 1 Hz vs 14.9 mJ (12.3-21.2) for 36 Hz (p = .003). Longer phase durations had no significant effect on discomfort despite generally requiring significantly lower current amplitudes. Phase durations 150, 200 and 400 µs required significantly lower energy consumption compared to 75 µs (all p ≤ .037). CONCLUSION: LI-NMES applied via a TTE sock produces a relevant plantar flexion of the ankle with the best comfort and lowest energy consumption using 1 Hz and phase durations 150, 200 or 400 µs.

Motor point heatmap guide for neuromuscular electrical stimulation of the quadriceps muscle.

PURPOSE: To create an anatomical chart that indicates the probability of finding a motor point (MP) in different areas of the quadriceps muscle. METHODS: On 31 healthy adults, the individual anatomy of the vastus medialis (VM), rectus femoris (RF) and vastus lateralis (VL) was determined using ultrasound. Thereafter, a 3 Hz neuromuscular electrical stimulation (NMES) MP-search with a MP-pen was performed. The thigh anatomy was normalized and divided into 112 (8x14) 3x3cm areas, and the probability of finding a MP in the different areas was calculated to create a MP heat-map. RESULTS: The heat-map displayed the two best 3x3cm areas, over VL and VM respectively, each with a probability greater than 50% of finding a MP and a higher probability compared to all other areas (p <.05). RF exhibited two areas with a 29% probability of finding a MP. A higher number of MPs on the quadriceps, mean (±SD) 9.4 ± 1, was in regression analysis found to be significantly associated with two independent factors higher physical activity level and lower body fat (R2 = 0.42, p=<.0001). CONCLUSION: Large inter-individual variations in location, and number of MPs were found, but the heat-map displayed areas with higher probability of finding a MP and can be used to facilitate NMES-application.

Knee extensor force production and discomfort during neuromuscular electrical stimulation of quadriceps with and without gluteal muscle co-stimulation.

PURPOSE: To investigate whether Neuromuscular Electrical Stimulation (NMES) simultaneously applied on the quadriceps (Q) and gluteal (G) muscles, as compared to single Q-stimulation alters the knee extensor force production and discomfort. METHODS: A total of 11 healthy participants (6 females), with normal weight and age between 19 and 54 years were included. The unilateral, isometric maximal voluntary contraction (MVC) was assessed for each participant in an isokinetic dynamometer (Biodex, system 3). NMES was, in a randomized order, applied only on the Q-muscle and on the Q- and G-muscles (QG) simultaneously. NMES-intensity was increased stepwise until the maximal tolerable level was reached regarding discomfort, graded according to the visual analogue scale (VAS). VAS and the % of MVC produced by NMES, were registered for each level, expressed as median (inter-quartile range). RESULTS: The maximum tolerated NMES-intensity applied on Q compared to QG resulted in equally high discomfort, 8.0 (6.0-9.0) vs 8.0 (6.3-9.0), and in equivalent knee extensor force production, 36.7 (29.9-47.5) and 36.2 (28.9-49.3), respectively, in % of MVC. At 20% of MVC, NMES applied on Q compared to QG resulted in equal acceptable discomfort, 3.0 (2.0-4.5) vs 3.0 (3-5.5), and comparable intensity levels, 41.5 (38.0-45.8) vs 43.5 (37.0-48.8), respectively. CONCLUSIONS: Simultaneous QG-NMES, as compared to single Q-NMES, does not seem to affect the knee extensor force production or discomfort. Q-NMES, without voluntary muscle contraction, can with an acceptable level of discomfort result in at least 20% of MVC.

Effects of electrode size and placement on comfort and efficiency during low-intensity neuromuscular electrical stimulation of quadriceps, hamstrings and gluteal muscles.

BACKGROUND: Neuromuscular electrical stimulation (NMES) may prevent muscle atrophy, accelerate rehabilitation and enhance blood circulation. Yet, one major drawback is that patient compliance is impeded by the discomfort experienced. It is well-known that the size and placement of electrodes affect the comfort and effect during high-intensity NMES. However, during low-intensity NMES the effects of electrode size/placement are mostly unknown. Therefore, the purpose of this study was to investigate how electrode size and pragmatic placement affect comfort and effect of low-intensity NMES in the thigh and gluteal muscles. METHODS: On 15 healthy participants, NMES-intensity (mA) was increased until visible muscle contraction, applied with three electrode sizes (2 × 2 cm, 5 × 5 cm, 5 × 9 cm), in three different configurations on quadriceps and hamstrings (short-transverse (ST), long-transverse (LT), longitudinal (L)) and two configurations on gluteus maximus (short-longitudinal (SL) and long-longitudinal (LL)). Current-density (mA/cm2) required for contraction was calculated for each electrode size. Comfort was assessed with a numerical rating scale (NRS, 0-10). Significance was set to p < 0.05 and values were expressed as median (inter-quartile range). RESULTS: On quadriceps the LT-placement exhibited significantly better comfort and lower current intensity than the ST- and L-placements. On hamstrings the L-placement resulted in the best comfort together with the lowest intensity. On gluteus maximus the LL-placement demonstrated better comfort and required less intensity than SL-placement. On all muscles, the 5 × 5 cm and 5 × 9 cm electrodes were significantly more comfortable and required less current-density for contraction than the 2 × 2 cm electrode. CONCLUSION: During low-intensity NMES-treatment, an optimized electrode size and practical placement on each individual muscle of quadriceps, hamstrings and gluteals is crucial for comfort and intensity needed for muscle contraction.

Achilles tendon ruptures during summer show the lowest incidence, but exhibit an increased risk of re-rupture.

PURPOSE: Achilles tendon rupture (ATR) is a common injury. The knowledge of seasonal factors´ impact is incomplete, but may provide means for preventive approaches for Achilles tendon related morbidity. The aim of this study was to investigate seasonal variations in ATR incidence in relation to injury mechanism, adverse events including risk of re-rupture, and patient-reported outcome in adults in Stockholm, Sweden. METHODS: In total, 349 patients with unilateral acute Achilles tendon rupture, prospectively treated with standardized surgical techniques, were retrospectively assessed. Date of injury was assigned to one of the four internationally defined meteorological seasons in the northern hemisphere. Injury mechanism and the rate of adverse events; deep venous thrombosis, infection and re-rupture in relation to per-operative complications. Patient-reported outcome at 1 year was assessed with the validated Achilles tendon Total Rupture Score. RESULTS: ATR incidence was significantly highest during winter and spring, and lowest during summer (p < 0.05). The most common sporting activities associated with ATR were badminton, floorball and soccer (> 50%). The rate of soccer-related ATR was highest during summer (p < 0.05). Patients sustaining an ATR during summer, compared to other seasons, exhibited more per-operative complications (p < 0.05), a significantly higher risk of re-rupture (p < 0.05) and a lower rate of good outcome (n.s.). The risk of other adverse events after ATR did not differ between the seasons. CONCLUSION: Winter and spring are the high risk seasons for sports-related ATR and the risk sports are badminton, soccer and floorball. The reason for the higher risk of re-rupture after ATR repair during summer should be further investigated. LEVEL OF EVIDENCE: III.

Microcirculation after Achilles tendon rupture correlates with functional and patient-reported outcomes.

Patients with acute Achilles tendon rupture (ATR) display an extended healing process with varying clinical outcome. Poor microcirculatory blood flow has been suggested to be a significant factor for the healing process. However, whether microcirculation may predict healing outcome has been mostly unknown. Therefore, we investigated whether blood flow in the Achilles tendon may be associated with patient-reported and functional outcomes after ATR. In vivo laser-Doppler flowmetry was used to assess microvascular blood flow bilateral in the Achilles tendons, during post-occlusive reactive hyperemia, of nine patients with acute total ATR at 2 weeks post-operatively. At 3 months post-operatively, patient-reported outcome was assessed using Achilles tendon Total Rupture Score (ATRS). At 1 year a uniform outcome score, Achilles Combined Outcome Score (ACOS), was obtained by combining validated, independent, patient-reported (ATRS), and functional outcome (heel-rise test) measures. An improved combined patient-reported and functional outcome, ACOS, at 1 year was significantly correlated with higher maximum blood flow (r=.777, P=.040) in the injured limb. Furthermore, enhanced patient-reported outcome, ATRS, at 3 months, was associated with an elevated ratio of maximum to resting blood flow (r=.809, P=.015) in the uninjured limb. Blood flow in early tendon healing is associated with long-term patient-reported and functional outcomes after ATR. The microcirculatory blood flow of both the healing and contralateral Achilles tendon seems to determine the healing potential after injury.

Ageing, deep vein thrombosis and male gender predict poor outcome after acute Achilles tendon rupture.

AIMS: Patients with an acute Achilles tendon rupture (ATR) take a long time to heal, have a high incidence of deep vein thrombosis (DVT) and widely variable functional outcomes. This variation in outcome may be explained by a lack of knowledge of adverse factors, and a subsequent shortage of appropriate interventions. PATIENTS AND METHODS: A total of 111 patients (95 men, 16 women; mean age 40.3, standard deviation 8.4) with an acute total ATR were prospectively assessed. At one year post-operatively a uniform outcome score, Achilles Combined Outcome Score (ACOS), was obtained by combining three validated, independent, outcome measures: Achilles tendon Total Rupture Score, heel-rise height test, and limb symmetry heel-rise height. Predictors of ACOS included treatment; gender; age; smoking; body mass index; time to surgery; physical activity level pre- and post-injury; symptoms; quality of life and incidence of DVT. RESULTS: There were three independent variables that correlated significantly with the dichotomised outcome score (ACOS), while there was no correlation with other factors. An age of less than 40 years old was the strongest independent predictor of a good outcome one year after ATR (odds ratio (OR) 0.20, 95% confidence interval (CI) 0.08 to 0.51), followed by female gender (OR) 4.18, 95% CI 1.01 to 17.24). Notably, patients who did not have a DVT while immobilised post-operatively had a better outcome (OR 0.31, 95% CI 0.12 to 0.80). CONCLUSION: Over the age of 40 years, male gender and having a DVT while immobilised are independent negative predictors of outcome in patients with an acute ATR. Cite this article: Bone Joint J 2016;98-B:1635-41.

Intermittent pneumatic compression reduces the risk of deep vein thrombosis during post-operative lower limb immobilisation: a prospective randomised trial of acute ruptures of the Achilles tendon.

Deep vein thrombosis is a common complication of immobilising the lower limb after surgery. We hypothesised that intermittent pneumatic compression (IPC) therapy in outpatients who had undergone surgical repair of acute ruptures of the Achilles tendon could reduce the incidence of this problem. A total of 150 patients who had undergone surgical repair of the Achilles tendon were randomised to either treatment with IPC for six hours per day (n = 74) under an orthosis or treatment as usual (n = 74) in a plaster cast without IPC. At two weeks post-operatively, the incidence of deep vein thrombosis was assessed using blinded, double-reported compression duplex ultrasound. At this point, IPC was discontinued and all patients were immobilised in an orthosis for a further four weeks. At six weeks post-operatively, a second compression duplex ultrasound scan was performed. At two weeks, the incidence of deep vein thrombosis was 21% in the treated group and 37% in the control group (p = 0.042). Age over 39 years was found to be a strong risk factor for deep vein thrombosis (odds ratio (OR) = 4.84, 95% confidence interval (CI) 2.14 to 10.96). Treatment with IPC, corrected for age differences between groups, reduced the risk of deep vein thrombosis at the two-week point (OR = 2.60; 95% CI 1.15 to 5.91; p =0.022). At six weeks, the incidence of deep vein thrombosis was 52% in the treated group and 48% in the control group (OR 0.94, 95% CI 0.49 to 1.83). IPC appears to be an effective method of reducing the risk of deep vein thrombosis in the early stages of post-operative immobilisation of outpatients. Further research is necessary to elucidate whether it can confer similar benefits over longer periods of immobilisation and in a more heterogeneous group of patients.

Chemokine expression of CCL2, CCL3, CCL5 and CXCL10 during early inflammatory tendon healing precedes nerve regeneration: an immunohistochemical study in the rat.

PURPOSE: Chemokines are major promoters of repair and may regulate nerve ingrowth that is essential in tendon healing. The purpose of this study was to assess the temporal occurrence of different chemokines during Achilles tendon healing in relation to sensory nerve regeneration. Chemokine presence in tendon healing has not been studied previously. METHODS: Chemokine expression, nerve regeneration, angiogenesis and inflammatory cell occurrence during healing of Achilles tendon rupture in the rat were studied by immunohistochemistry and histology including semiquantitative assessment. Markers for chemokines (CCL5, CCL2, CCL3, CXCL10), nerves (PGP-9.5) and sensory neuropeptide substance P (SP) were analysed at different time points (1 day-16 weeks) post-rupture. RESULTS: In intact tendons (controls) immunoreactivity to all chemokines, PGP-9.5 and SP were confined to the tendon surroundings. After rupture, there was rapid increase in the tendon proper of the chemokines studied, all exhibiting their peak expression at week 1. Subsequently, at weeks 2-6, emerging inflammatory cells and maximum sprouting of PGP-/SP-positive nerves were observed close to newly formed blood vessels within the tendon proper, while chemokine expression already decreased. During weeks 6-8, PGP-/SP-positive nerves withdrew from the rupture site and relocated together with the chemokines in the surrounding tendon. CONCLUSIONS: Early chemokine expression in the healing tendon precedes ingrowth of new nerves, angiogenesis and emergence of inflammatory cells. The fine-tuned temporal and spatial appearance of chemokines suggests a chemoattractant role for inflammatory cell migration and possibly also a role in angiogenesis and neurogenesis. Chemokines may thus exhibit vital targets for biological modulation of tendon repair.

Can foot compression under a plaster cast prevent deep-vein thrombosis during lower limb immobilisation?

We hypothesised that adjuvant intermittent pneumatic compression (IPC) beneath a plaster cast would reduce the risk of deep-vein thrombosis (DVT) during post-operative immobilisation of the lower limb. Of 87 patients with acute tendo Achillis (TA) rupture, 26 were prospectively randomised post-operatively after open TA repair. The treatment group (n = 14) received two weeks of IPC of the foot for at least six hours daily under a plaster cast. The control group (n = 12) had no additional treatment. At two weeks post-operatively all patients received an orthosis until follow-up at six weeks. At two and six weeks the incidence of DVT was assessed using colour duplex sonography by two ultrasonographers blinded to the treatment. Two patients withdrew from the study due to inability to tolerate IPC treatment. An interim analysis demonstrated a high incidence of DVT in both the IPC group (9 of 12, 75%) and the controls (6 of 12, 50%) (p = 0.18). No significant differences in incidence were detected at two (p = 0.33) or six weeks (p = 0.08) post-operatively. Malfunction of the IPC leading to a second plaster cast was found to correlate with an increased DVT risk at two weeks (ϕ = 0.71; p = 0.019), leading to a premature abandonment of the study. We cannot recommend adjuvant treatment with foot IPC under a plaster cast for outpatient DVT prevention during post-operative immobilisation, owing to a high incidence of DVT related to malfunctioning of this type of IPC application.

Anti-inflammatory cytokine profile in early human tendon repair.

PURPOSE: The aim of this study was to assess inflammation and the presence and relative levels of cytokines, which may be involved in regulating early human Achilles tendon healing. METHODS: Nine patients with acute Achilles tendon rupture were included, operated on and post-operatively immobilized. Two weeks post-operatively, microdialysis of the peritendinous interstitial compartment was performed in the healing and intact contralateral Achilles tendons. Quantification of tumour necrosis factor (TNF)-α, interferon (IFN)-γ, interleukin (IL)-1ß, IL-6, IL-8, IL-10, IL-12p70 and IL-17A was accomplished using a cytometric bead array. Prostaglandin (PG) E2 levels were measured by enzyme immunoassay. RESULTS: None of the patients displayed detectable PGE2 levels. Pro-inflammatory cytokines were below detection levels (IFNγ, IL-12, and IL-17) or did not differ between injured and control tendons (IL-1ß and TNF). Notably, IL-6, IL-8 and IL-10 concentrations in the healing Achilles tendon were significantly elevated: 13-fold (p = 0.009), 28-fold (p = 0.02), and 3.7-fold (p = 0.03), respectively. CONCLUSION: At 2 weeks post-human Achilles tendon rupture, healing is characterized by a resolving inflammatory phase and up-regulation of IL-6, IL-8 and IL-10. The absence of inflammation suggests that at this time point, these cytokines may be associated with anti-inflammatory and regenerative effects on the tendon healing process.

Metabolic activity in early tendon repair can be enhanced by intermittent pneumatic compression.

Since Achilles tendon healing is protracted, more knowledge of metabolites known to meet the demands for biosynthesis and proliferation is needed. We hypothesized that essential metabolites, glutamate, glucose, lactate, pyruvate and glycerol, are present and upregulated in healing Achilles tendons. We moreover hypothesized that adjuvant intermittent pneumatic compression (IPC), which increases blood flow, upregulates metabolite concentrations. Twenty patients with acute Achilles tendon rupture were recruited, operated, and included. The control group, 15 patients, received plaster cast immobilization, while five patients received adjuvant foot IPC beneath the plaster cast. At 2 weeks postoperatively, microdialysis of the healing and contralateral intact Achilles tendons was followed by quantification of metabolites. Healing compared to intact tendons of the controls exhibited significantly increased concentrations (mM) of glutamate (60 ± 14 vs 20 ± 11), lactate (1.15 ± 0.60 vs 0.64 ± 0.35), and pyruvate (81 ± 29 vs 35 ± 25, µM). Healing tendons of the IPC vs control group displayed higher levels of glutamate (84 ± 15 vs 62 ± 16) and glucose (3.44 ± 0.62 vs 2.62 ± 0.72); (P < 0.05) and trends toward higher concentrations of pyruvate, lactate, and glycerol (P < 0.10). The present study demonstrates that early Achilles tendon repair entails and upregulates local essential metabolites. This metabolic response can, during tendon healing with plaster cast immobilization, be promoted by adjuvant IPC.

Substance P injections enhance tissue proliferation and regulate sensory nerve ingrowth in rat tendon repair.

Tendon healing is characterized mostly by slow rehabilitation and, as in tendinopathy, aberrant, protracted sensory nerve ingrowth. This study investigated whether administration of the sensory neuropeptide substance P (SP) could enhance healing and modulate sensory nerve plasticity after Achilles tendon rupture. Fifty-four male Sprague-Dawley rats were allocated to three groups, all receiving six daily injections post-rupture of; (1) SP (10(-6) mol/kg body weight)+endopeptidase inhibitors captopril and thiorphan, (2) captopril/thiorphan only and (3) saline control. At 1, 3 and 6 weeks post-rupture tendon healing was evaluated by assessments of fibroblast proliferation, collagen III-LI (like) occurrence, diameter of newly organized collagen and sensory nerve fiber ingrowth. At 1 week, the SP-treated group exhibited increased occurrence of collagen III-LI (P=0.03) and of organized collagen (P=0.04) compared with control. At 3 weeks, the SP group notably displayed reduced SP-nerve fiber ingrowth (P=0.02), and higher fibroblast density (P=0.004). Both the SP and captopril/thiorphan groups demonstrated increase in collagen fiber organization compared with control (P=0.02 and 0.004, respectively). At 6 weeks, no significant differences were observed between the groups. SP supply in tendon repair promotes early tissue proliferation and regulation of endogenous sensory nerve ingrowth, suggesting implications for novel treatment in tendinopathy.

Coexistence of up-regulated NMDA receptor 1 and glutamate on nerves, vessels and transformed tenocytes in tendinopathy.

Elevated levels of the neurotransmitter glutamate and the presence of its receptor, N-methyl-d-aspartate receptor type 1 (NMDAR1), have been established in patients with tendinopathy, i.e. chronic tendon pain and degeneration. However, whether NMDAR1 is up- or down-regulated in tendinopathy and co-localized with glutamate is still unexplored. We hypothesize that an alteration in tissue expression and in the coexistence of NMDAR1 and glutamate occurs in tendinopathy and might play a role in nociception and possibly also progression of tendon degeneration (tendinosis). We therefore examined the tissue distribution and levels of NMDAR1 and glutamate in biopsies from patients with patellar tendinopathy (n=10) and from controls (n=8). The biopsies were single- and double-stained immunohistochemically for glutamate and NMDAR1 and assessed subjectively and semi-quantitatively. The chronic painful tendons exhibited a significant elevation of NMDAR1 (ninefold), which was independent of the observed increase in glutamate (10-fold). This up-regulation of NMDAR1 and glutamate was found to be co-localized on nerve fibers as well as on morphologically altered tenocytes and blood vessels. None of the controls exhibited neuronal coexistence of glutamate and NMDAR1. The neuronal coexistence of glutamate and NMDAR1, observed in painful tendinosis but not in controls, suggests a regulatory role in intensified pain signalling.

Status of the PCDD and PCDF contamination of commercial milk caused by milk cartons.

Within the scope of this study, possible migration of polychlorinated dibenzodioxins and -furans (PCDDs/PCDFs) from cartons (produced using bleached cardboard) into cow's milk was investigated. Three different types of carton were examined. The milk samples were taken at different times of storage and analysed for PCDDs/PCDFs. In contrast to a similar study carried out in 1990, the I-TEQ-results of all cartons analysed may be considered very low. No significant migration of toxic (2,3,7,8-chlorine-substituted) PCDDs/PCDFs could be observed. This is due to the very low PCDD/PCDF concentrations in modern cardboards as a result of the elemental chlorine-free (ECF) bleaching processes used. As far as the milk samples are concerned, no influence of the cardboards (according to the I-TEQ data) could be ascertained. I-TEQ concentrations in milk did not noticeably change during storage times of three, six and eight days. The I-TEQ-data obtained for all milk samples was found to be in the range typical of background concentrations in cow's milk in Germany. Although 1,2,7,8-TCDF is not included in the calculation of the TEQ (no 2,3,7,8-chlorine-substitution), analysis of this congener in cardboard samples was also carried out as a matter of general interest for cardboard investigations and an indicator of pulp bleaching with free chlorine.

An opioid system in connective tissue: a study of achilles tendon in the rat.

The occurrence of endogenous opioids and their receptors in rat achilles tendon was analyzed by immunohistochemistry (IHC), radioimmunoassay (RIA), and in vitro binding assays. The investigation focused on four enkephalins, dynorphin B, and nociceptin/orphanin FQ. Nerve fibers immunoreactive to all enkephalins (Met-enkephalin, Leu-enkephalin, Met-enkephalin-Arg-Gly-Lys, Met-enkephalin-Arg-Phe) were consistently found in the loose connective tissue and the paratenon, whereas dynorphin B and nociceptin/orphanin FQ could not be detected. The majority of enkephalin-positive nerve fibers exhibited varicosities predominantly seen in blood vessel walls. Measurable levels of Met-enkephalin-Arg-Phe and nociceptin/orphanin FQ were found in tendon tissue using RIA, whereas dynorphin B could not be detected. In addition to the endogenous opioids identified, delta-opioid receptors on nerve fibers were also detected by IHC. Binding assays to characterize the opioid binding sites showed that they were specific and saturable for [3H]-naloxone (Kd 7.01 +/- 0.98 nM; Bmax 23.52 +/- 2.23 fmol/mg protein). Our study demonstrates the occurrence of an opioid system in rat achilles tendon, which may be assumed to be present also in other connective tissues of the locomotor apparatus. This system may prove to be a useful target for pharmacological therapy in painful and inflammatory conditions by new drugs acting selectively in the periphery.

Autonomic innervation of tendons, ligaments and joint capsules. A morphologic and quantitative study in the rat.

We analyzed the neuronal occurrence of autonomic transmitters; noradrenaline (NA), neuropeptide Y (NPY) and vasoactive intestinal polypeptide (VIP), in the Achilles tendon, medial and lateral collateral ligaments and knee joint capsule in the rat--by immunohistochemistry (IHC). In addition, the tissue concentrations of the sympathetic neuropeptide, NPY, and the parasympathetic peptide, VIP, were determined by radioimmunoassay (RIA). IHC demonstrated nerve fibers containing sympathetic vasoconstrictors--NA and NPY--and the parasympathetic vasodilator, VIP, in all tissues. NPY- and NA-positive nerve fibers were predominantly observed in larger blood vessels, whereas, nerve fibers immunoreactive to VIP were found in smaller vessels. In many nerve fibers a co-localization of the transmitters was seen. RIA showed that the concentration of NPY compared to VIP was 15-times higher in ligaments and twice as high in tendons and capsules. The differences noted may reflect a difference in vulnerability to degenerative conditions. In pathological conditions, dysregulation of autonomic transmitters in hypovascularized tissues subjected to repetitive mechanical load may contribute to tissue hypoxia leading to degeneration and rupture of tendons and ligaments.