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PubMed1990

Mechanical function of the human lumbar interspinous and supraspinous ligaments.

The mechanical function of the interspinous and supraspinous ligaments has been examined by simulating, on excised specimens, the deformation caused during forward flexion of the spine in real life. The load extension curves showed that during the first half of flexion the ligaments carried very little load but towards the end of the range of flexion they resisted up to 134N. When the supraspinous ligament was sectioned the interspinous ligament alone resisted 75% of this load. Testing at high strain rates showed an increase in load-carrying capacity of up to 30%. The maximum extension moment that can be produced by these ligaments was calculated to be approximately 7 Nm, or 5% of the moment the back muscles can produce across any intervertebral joint. Hence, during active lifting, these ligaments in isolation provide little mechanical assistance.

Aged↗
PubMed1990

Immunocytochemical localization of phosphatidylinositol-anchored acetylcholinesterase in excitable membranes of Torpedo ocellata.

In Torpedo electric organ much of the acetylcholinesterase is a 'globular' dimer (G2), anchored to the plasma membrane via covalently attached phosphatidylinositol and solubilized by a bacterial phosphatidylinositol-specific phospholipase C. This suggested that selective solubilization with phosphatidylinositol-specific phospholipase C, coupled with immunocytochemistry, might be used to localize G2 acetylcholinesterase in excitable tissues of Torpedo. Cryostat sections of electric organ, electromotor nerve, electric lobe and back muscle from Torpedo ocellata were labelled, using three different antibody preparations to Torpedo acetylcholinesterase, followed by a fluorescent second antibody, before and after exposure to the phospholipase. Sites of innervation on electrocytes and myofibers were labelled selectively, as were motor and electromotor nerves. In all these cases labelling was substantially diminished by prior exposure to the phospholipase. The results support our previous assignment, based on biochemical evidence, for a neuronal and synaptic localization of the G2 acetylcholinesterase in Torpedo. Electric lobe acetylcholinesterase appears insensitive to the phospholipase treatment and lacks certain epitopes present in both electric organ and electromotor nerve enzyme. This suggests that substantial processing of the G2 form occurs concomitantly with its movement from the electric lobe into the electromotor nerve.

Acetylcholinesterase↗
PubMed1996

Thyrotropin-releasing hormone (TRH) has independent excitatory and modulatory actions on lamina IX neurons of lumbosacral spinal cord slices from adult rats.

Coronal and horizontal slices of the lumbar and sacral spinal cord, respectively, of ovariectomized adult rats, either treated with estrogen (OVX+E) or untreated (OVX), were used to test the neuronal actions of TRH and its metabolite, cyclo(His-Pro) (or cHP). Both coronal slices, which possess only short stumps of ventral roots (VRs), and horizontal slices, in which long sections of VRs were preserved, were used for extracellular recording of single motor and other types of neurons. Methodological comparisons between these two types of slices showed that the length of VRs preserved had no significant effect on the characteristics of motoneurons (MNs). In coronal slices, MNs in medial and lateral lamina IX (MNM and MNL, respectively) were identified by antidromic activation. Of these lumbar MNs, estrogen treatment lowered the antidromic activation threshold for MNM but not MNL. Because MNM innervate the back muscles crucial for the execution of the estrogen-dependent lordosis, the observed estrogen effect may contribute to the hormone's induction of the sexual behavior. The recorded MNs and other types of neurons were subjected to bath applications of TRH, cHP, and neurotransmitters. TRH was found to be capable of evoking an early, shorter-lasting neuronal excitation and/or a late, longer-lasting modulation of neuronal responses to transmitters. Each neuronal action could occur with or without the other, and the occurrence of the excitation did not affect the probability of whether a modulation would occur later. The modulatory, but not the excitatory, action appeared to be shared by cHP, because cHP could also modulate neuronal responses in similar, if not identical, ways as TRH did, but could neither stimulate neurons nor mimic TRH in desensitizing TRH-evoked excitation. The modulatory actions of the two peptides were not affected by estrogen. Although the excitatory action was desensitized by repeated TRH applications, the modulatory action did not appear to be attenuated but instead was often enhanced by repeated administrations of TRH and/or cHP. These results, together with the essentially identical findings from our previous study on hypothalamic neurons, indicate that the excitatory and the modulatory actions of TRH are independent of each other and, hence, are mediated by different subcellular mechanisms.

Animals↗
PubMed1990

The pattern of axonal degeneration in the peripheral nervous system varies with different types of lesion.

Degeneration of axons in the mouse sciatic nerve was examined using conventional silver staining and by noting the presence or absence of a compound action potential on stimulating the nerve distal to the point of crush or cut. The presence of myeloperoxidase-positive cells was also examined in frozen sections of the nerve. In all experiments the distal, disconnected segment was studied. Degeneration after crushing with fine watchmakers forceps always began in the most distal part of the nerve and proceeded in a distoproximal direction, from the nerve entry point into a muscle back to the crush site. Myeloperoxidase-positive cells were also recruited into the nerve starting at the distal rather than proximal (the originally injured) end. This result favours the view that degeneration is triggered by lack of trophic support from the cell body rather than entry of deleterious substances at the site of the injury, for the terminals furthest from both the source of supply and the injury are affected first. Degeneration following nerve section was always more rapid than after crushing. The rate of axonal sealing at the injury was, however, no slower than after crushing, so it does not seem likely that greater entry of possible degeneration-triggering material at the injury site is the explanation for this. Crush lesions in which the perineurium was also cut open and the blood supply at the site was damaged, degenerated at the slower rate characteristic of simple crushes.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1986

Acid solutions enhance bone-inducing activity of a murine osteosarcoma.

Heterotopic osteogenesis induced by lyophilized powder of a murine osteosarcoma was greatly enhanced by treating the powder with the following acids: 0.01 N HCl (pH 2.0), 0.1 N HCl (pH 1.7), 1 N HCl (pH 1.2), 0.5 M acetic acid (pH 2.8), 0.5 M lactic acid (pH 2.0), and 0.5 M citric acid (pH 1.8). The acid-treated preparations were neutralized before implantation into back muscles of ddY mice. Acid treatment resulted in almost a fivefold increase in the amount of new bone formation, as measured by uptake of strontium-85 into the implants. In addition bone-inducing activity was increased by longer exposure to 0.01 N HCl up to 24 h. These findings suggest that the bone-inducing activity of the osteosarcoma is enhanced by various acids below pH 2.8. A possible role for hydrogen ion concentration in the regulation of osteogenesis is discussed.

Acids↗
PubMed1989

Establishment of an osteoinductive murine osteosarcoma clonal cell line showing osteoblastic phenotypic traits.

A clonal cell line, BFO-6, was established in culture from a Dunn osteosarcoma cell line (BFO) and characterized on the basis of bone-inducing activity, alkaline phosphatase activity, PTH sensitive cAMP production and tumorigenicity. Lyophilized pellets of devitalized cells, when implanted into the back muscles of allogenic host mice, consistently elicited heterotopic bone formation at 3 weeks postimplantation. BFO-6 cells were found to be rich in alkaline phosphatase activity and showed a significant response to h-PTH stimulation. The doubling time in the logarithmic phase was 17.2 h and the cells revealed an acrocentric karyotype. Subcutaneous transplantation of cells (1 x 10(7) cells) into a C3H mouse resulted in the production of a tumor with histological features of osteosarcoma. This tumor also retained bone-inducing activity and high alkaline phosphatase activity.

Alkaline Phosphatase↗
PubMed1990

Matrix vesicle calcification of ectopically induced osteoid tissue in 1-hydroxethylidene-1,1-bisphosphonate (HEBP)-treated mice.

The effects of 1 hydroxethylidene-1, 1-bisphosphonate (HEBP) on the mineralization of osteoid induced ectopically by a murine, osteosarcoma-derived, bone-inducing substance was examined at the ultrastructural level. Samples of Dunn osteosarcoma were chemically processed, lyophilized into pellets of uniform dimensions and protein content, and implanted into the back muscles of male, 4 week old, ddY mice. In control animals injected intraperitoneally (i.p.) with saline each day for 30 days, the pellets exhibited ectopic cartilage and bone formation in a sequential manner consistent with endochondral ossification. Daily administration of HEBP (30 mg/kg i.p. per day) to animals over the same period produced wide anastomosing strands of osteoid and haematopoietic marrow in the pellets. Electron microscopical examination revealed a tightly packed organization of collagen fibrils with many membrane bounded organelles or matrix vesicles (0.05-0.3 micron diameter) dispersed throughout this nonmineralized matrix. After withdrawal of HEBP, mineralization proceeded initially within matrix vesicles and secondarily by involvement of the surrounding collagen fibrils. In pellets retrieved at 30 days after HEBP withdrawal, most of the matrix had been mineralized and remodeled to reveal thin bony trabeculae and an overall histologic appearance similar to that seen in the control pellets. This model of ectopic bone formation permits clear visualization of the sequence of mineral nucleating events in osteoid in the absence of an established mineralizing front. The results add further support to the hypothesis that matrix vesicles are directly responsible for the initiation of mineralization in bone.

Animals↗
PubMed2004

A prostanoid receptor EP4 agonist enhances ectopic bone formation induced by recombinant human bone morphogenetic protein-2.

The anabolic effects of prostaglandin E(2) on bone are effected through the activation of EP4, a G protein-coupled receptor. In the present study, we examined the effects of a prostanoid receptor-selective agonist (ONO-4819) in an experimental system of ectopic bone formation using recombinant human bone morphogenetic protein-2 (rhBMP-2). Collagen pellets containing rhBMP-2 were implanted onto the back muscles of mice and then treated with ONO-4819 administered every 8 h by subcutaneous injection. The ossicles elicited ectopically by rhBMP-2 in mice treated with 30 microg/kg ONO-4819 were significantly larger in size and had a higher bone mineral density and bone mineral content when compared to the controls. We also noted that the anabolic effect of ONO-4819 was seen only in the early phase of the rhBMP-2-induced bone-forming process. These experimental results indicate that the EP4 receptor agonist enhances the rhBMP-2-induced bone formation through a selective effect on early stage mesenchymal cells, which in turn may result in increased responsiveness of the host animals to rhBMP-2.

Absorbable Implants↗
PubMed2005

Biological performance of uncoated and octacalcium phosphate-coated Ti6Al4V.

The in vivo behavior of a porous Ti6Al4V material that was produced by a positive replica technique, with and without an octacalcium phosphate (OCP) coating, has been studied both in the back muscle and femur of goats. Macro- and microporous biphasic calcium phosphate (BCP) ceramic, known to be both osteoconductive and able to induce ectopic bone formation, was used for comparison purpose. The three groups of materials (Ti6Al4V, OCP Ti6Al4V and BCP) were implanted transcortically and intramuscularly for 6 and 12 weeks in 10 adult Dutch milk goats in order to study their osteointegration and osteoinductive potential. In femoral defects, both OCP Ti6Al4V and BCP were performing better than the uncoated Ti6Al4V, at both time points. BCP showed a higher bone amount than OCP Ti6Al4V after 6 weeks of implantation, while after 12 weeks, this difference was no longer significant. Ectopic bone formation was found in both OCP Ti6Al4V and BCP implants after 6 and 12 weeks. The quantity of ectopically formed bone was limited as was the amount of animals in which the bone was observed. Ectopic bone formation was not found in uncoated titanium alloy implants, suggesting that the presence of calcium phosphate (CaP) is important for bone induction. This study showed that CaPs in the form of coating on metal implants or in the form of bulk ceramic have a significantly positive effect on the bone healing process.

Alloys↗
PubMed2005

3D microenvironment as essential element for osteoinduction by biomaterials.

In order to unravel the mechanism of osteoinduction by biomaterials, in this study we investigated the influence of the specific surface area on osteoinductive properties of two types of calcium phosphate ceramics. Different surface areas of the ceramics were obtained by varying their sintering temperatures. Hydroxyapatite (HA) ceramic was sintered at 1150 and 1250 degrees C. Biphasic calcium phosphate (BCP) ceramic, consisting of HA and beta-tricalcium phosphate (beta-TCP), was sintered at 1100, 1150 and 1200 degrees C. Changes in sintering temperature did not influence the chemistry of the ceramics; HA remained pure after sintering at different temperatures and the weight ratio of HA and beta-TCP in the BCP was independent of the temperature as well. Similarly, macroporosity of the ceramics was unaffected by the changes of the sintering temperature. However, microporosity (pore diameter <10 microm) significantly decreased with increasing sintering temperature. In addition to the decrease of the microporosity, the crystal size increased with increasing sintering temperature. These two effects resulted in a significant decrease of the specific surface area of the ceramics with increasing sintering temperatures. Samples of HA1150, HA1250, BCP1100, BCP1150 and BCP1200 were implanted in the back muscles of Dutch milk goats and harvested at 6 and 12 weeks post implantation. After explantation, histomorphometrical analysis was performed on all implants. All implanted materials except HA1250 induced bone. However, large variations in the amounts of induced bone were observed between different materials and between individual animals. Histomorphometrical results showed that the presence of micropores within macropore walls is necessary to make a material osteoinductive. We postulate that introduction of microporosity within macropores, and consequent increase of the specific surface area, affects the interface dynamics of the ceramic in such a way that relevant cells are triggered to differentiate into the osteogenic lineage.

Animals↗
PubMed2005

Low dose fibroblast growth factor-2 (FGF-2) enhances bone morphogenetic protein-2 (BMP-2)-induced ectopic bone formation in mice.

To examine how fibroblast growth factor-2 (FGF-2) affects the BMP signaling pathway during bone morphogenetic protein-2 (BMP-2)-induced ectopic bone formation, we implanted type I collagen disks containing constant amounts of BMP-2 (5 micrograms) and varying amounts of FGF-2 onto the back muscles of adult male mice. We then performed histological analyses and histomorphometry, and measured bone mineral density and radiopaque area on the discs 1, 2, and 3 weeks after implantation. We also determined the expression profiles of several genes involved in bone formation and the BMP signaling pathway in the muscle that had been adjacent to the implanted disc and in muscle-derived primary culture cells that had similarly been treated with a constant concentration of BMP-2 and a varying concentration of FGF-2. In the presence of a constant amount of BMP-2, we confirmed that low doses of FGF-2 increased ectopic bone formation in vivo and high doses inhibited bone formation. Northern and/or Western blots of recovered muscle from the in vivo experiment and treated muscle-derived primary culture cells from the in vitro experiment revealed that low doses of FGF-2, but not high doses, increased the expression BMP receptor (BMPR)-1B, phosphorylated Smad1, Noggin, and Osteocalcin. Our results indicate that low-dose FGF-2 may facilitate BMP-2-induced ectopic bone formation by altering the expression of BMPRs on the surface of bone forming progenitor cells. They also indicate that the inhibitory effect of high-dose FGF-2 is not mediated via increased expression of the BMP inhibitor Noggin.

Animals↗
PubMed2004

Decreased social behaviour following 3,4-methylenedioxymethamphetamine (MDMA) is accompanied by changes in 5-HT2A receptor responsivity.

This study examined the involvement of the 5-HT(2A) receptor in the long-term anxiogenic effect of a brief exposure of young rats to 3,4-methylenedioxymethamphetamine (MDMA) using the social interaction and elevated plus-maze paradigms. Wistar rats (post-natal day (PND) 28) received either MDMA (5 mg/kg i.p.) or saline (1 ml/kg i.p.) hourly for 4 h on 2 consecutive days. Locomotor activity was measured for 60 min after the first injection and core body temperature was recorded at regular intervals over 4 h. On PND 84, without further drug administration, social interaction was assessed between treatment-matched rat pairs derived from separate litters. On PND 86, rats received either the 5-HT(2A/2C) receptor agonist, 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI, 1 mg/kg i.p.) or saline and locomotor activity, wet-dog shakes and back muscle contractions were monitored. The change in elevated plus-maze behaviour was assessed following the same injection on PND 87. Acutely, MDMA produced a significant hyperlocomotion and hyperthermia (p<0.01). Following 55 days of abstinence, social interaction was reduced by 27% in MDMA pre-treated rats compared with that in controls (p<0.01). On the elevated plus-maze, pre-treatment with MDMA prevented the anxiogenic effect of DOI. On PND 92, hippocampal, frontal cortical and striatal 5-hydroxytryptamine (5-HT) was significantly reduced in MDMA pre-treated rats by between 16% and 22%, without any accompanying change in [(3)H]paroxetine binding in cortical homogenates. In conclusion, exposure of young rats to repeated MDMA caused serotonin depletion and induced 'anxiety-like' behaviour in the social interaction test accompanied by a long-lasting reduction in specific 5-HT(2A) receptor mediated behaviour.

Amphetamines↗
PubMed1994

Kinetic mechanism of myofibril ATPase.

The kinetic mechanism of myofibril ATPase was investigated using psoas and mixed back muscle over a range of ionic strengths. Myofibrils were labeled with pyrene iodoacetamide to measure the rate constants for the binding of ATP and formation of the weakly attached state. The velocity of shortening was measured by stopping the contraction at various times by mixing with pH 4.5 buffer. The transient and steady-state rates of ATP hydrolysis were measured by the quench flow method. The results fitted the kinetic scheme [formula: see text] The rate constants (or equilibrium constants for steps 1 and 6) were obtained for the six steps. k5 was calculated from the KM for shortening velocity, K1, and k2. The rate constants were essentially equal for myofibrils and acto-S-1 at low ionic strength. Increasing the ionic strength up to 100 mM in NaCl increased the rate of the hydrolysis step and the size of the phosphate burst and the effective rate of product release became the rate-limiting step. The step calculated from the velocity of shortening, k5, and k2 is 15 nm, based on a model in which step 4 is the force-generating step.

Adenosine Diphosphate↗
PubMed1987

Differential susceptibility to phosphatidylinositol-specific phospholipase C of acetylcholinesterase in excitable tissues of embryonic and adult Torpedo ocellata.

The ability of phosphatidylinositol-specific phospholipase C (PIPLC) to solubilize acetylcholinesterase (AChE) in the electromotor system of adult Torpedo ocellata and in the developing electric organ was examined. PIPLC solubilizes significant amounts of the membrane-bound G2 form of AChE throughout embryonic development of the electric organ, as it does in the adult electric organ, the AChE of which we have shown to contain covalently bound inositol in its membrane-anchoring domain. In the electromotor system of the mature fish, PIPLC solubilizes almost quantitatively the AChE dimer in the electromotor axon as in the electric organ itself, but the corresponding fraction in the electric lobe is almost totally resistant to the phospholipase. This finding implies that the covalently bound phosphatidylinositol is added concomitantly with axonal transport. A substantial part of the G2 form in back muscle is sensitive to PIPLC, whereas the G4 tetramer of Torpedo brain is completely resistant.

Acetylcholinesterase↗
PubMed1990

Augmentation myoperitoneocystoplasty.

The use of isolated intestinal segments is currently the best method of augmenting bladder capacity. The ability to perform a successful augmentation cystoplasty using nonintestinal tissue would decrease the morbidity of the operation. We studied the use of skeletal muscle-backed parietal peritoneal tissue in augmentation cystoplasty. In the experimental group animals, bladder capacity increased by 67% and urothelial regeneration was observed histologically. Problems with electrolyte abnormalities, urosepsis, or excessive mucous production were not encountered. In conclusion, a myoperitoneal flap may serve as a template for augmentation cystoplasty. Further investigation using this technique is warranted.

Animals↗
PubMed1999

Modification of 5-HT2 receptor mediated behaviour in the rat by oleamide and the role of cannabinoid receptors.

Oleamide (cis-9,10-octadecenoamide) is an endogenous brain lipid which has been suggested to induce sleep in experimental animals. The mechanism of action is unclear but shares many of the characteristics of endogenous cannabinoids such as anandamide and has been shown to enhance in vitro responses to 5-HT and GABA. In the present study we investigated the effects of oleamide on two motor behaviours, back muscle contractions (BMC) and wet-dog shakes (WDS) induced in rats by treatment with the 5-HT2 receptor agonist DOI ((+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane hydrochloride). We then examined the potential involvement of CB1 cannabinoid receptors in the responses to oleamide and the mechanism of interaction between CB1 and 5-HT2 receptors. Oleamide and the cannabinoid receptor agonist HU210 (6aR)-trans-3-(1,1-dimethylheptyl)6a,7,10,10a-tetrahydro-1-h ydroxy-6,6-dimethyl-6H-dibenzo[b,d]pyran-9-methanol) produced a hypolocomotion which was prevented by the CB1 antagonist SR141716A (N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-me thyl-1H-pyrazole-3-carboxamide hydrochloride). Despite having no effect alone, oleamide and HU210 potentiated BMC induced by treatment with DOI. SR141716A alone did not affect the response to DOI but it blocked the potentiations caused by oleamide or HU210. WDS were unaffected by oleamide and slightly reduced by HU210. In vitro, oleamide and HU210 enhanced the high affinity binding of 5-HT to 5-HT2 receptors on rat cerebral cortex membranes labelled with 3H-ketanserin. Neither agent, however, altered 5-HT-stimulated phosphoinositide hydrolysis in rat cerebral cortex slices. Oleamide occupied CB1 cannabinoid receptors on rat brain membranes labelled with 3H-CP55940 with an IC50 of 10 microM. The data presented are consistent with oleamide acting via a cannabinoid recognition site to enhance 5-HT2 receptor function in vivo. The mechanism of the modulation is still unclear but it does not appear to involve a potentiation of 5-HT2 receptor-stimulated phosphoinositide hydrolysis.

Amphetamines↗
PubMed1998

Rabbit urethral regeneration using small intestinal submucosa onlay grafts.

OBJECTIVES: To determine if small intestinal submucosa (SIS) can evoke urethral regeneration. METHODS: Twenty male white New Zealand rabbits were assigned to one of three experimental groups. Group 1 (n = 4) underwent simple urethrotomy and closure. Group 2 (n = 8), a second control group, underwent an onlay urethroplasty with a graft of full-thickness preputial skin from the host rabbit. Group 3 (n = 8) underwent an onlay urethroplasty with an SIS graft. RESULTS: All eight SIS onlay grafts promoted regeneration of the normal rabbit epithelium supported by a well-vascularized collagen and smooth muscle backing. Preputial free onlay grafts maintained a keratinizing squamous cell epithelium with a poor supportive backing, which resulted in the formation of urethral diverticulum. CONCLUSIONS: SIS onlay patch grafts for urethroplasty promote rabbit urethral regeneration.

Animals↗
PubMed2002

Alteration in 5-hydroxytryptamine agonist-induced behaviour following a corticosterone implant in adult rats.

Hypercortisolism and altered serotonergic function may account for the pathological symptoms seen in depression. This study examines the impact of 4 days continuous corticosterone treatment on 5-HT agonist-induced behaviour to delineate changes in 5-HT receptor function in the adult rat. The flat body posture, reciprocal forepaw treading, elevated corticosterone, hyperglycaemia, hypothermia and reduced hippocampal 5-HT induced by the 5-HT(1A) agonist 8-OHDPAT (0.3 mg/kg ip) were all significantly attenuated by the corticosterone implant. The elevation in plasma corticosterone and back muscle contractions evoked by the 5-HT(2A) agonist DOI (1 mg/kg ip) were attenuated, whilst wet-dog shakes were enhanced by corticosterone treatment. 5-HT(2B) agonist-induced behaviour and the hypolocomotion and hypophagia induced by the 5-HT(2C) agonist m-CPP (2.5 mg/kg ip) were unaltered but the mCPP-induced elevation in corticosterone was abolished by corticosterone treatment. Hypothalamic 5-HT receptors mediating corticosterone- and 5-HT(1A) receptors, whether on serotonergic nerve terminals or postsynaptic neurones, were downregulated by corticosterone treatment. In contrast, 5-HT(2A) receptors may be up- or downregulated dependent on whether they are on supraspinal or spinal neurones, respectively. A comparison of the brain region-dependent alteration in serotonergic function produced by hypercorticosterone in the rat with that seen in depression is discussed.

Amphetamines↗