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B Heppelmann

Publications and source records attributed to B Heppelmann.

At least 37 records · Page 2Linked to original sources
PubMed1997

Quantitative examination of calcitonin gene-related peptide immunoreactive nerve fibres in the cat knee joint capsule.

The knee joint of the cat has been used extensively to study the morphology and function of primary afferents in a deep somatic tissue. A proportion of these neurones synthesizes various neuropeptides, with calcitonin gene-related peptide being the most prominent. In the present study we examined the distribution and density of nerve fibers immunoreactive for calcitonin gene-related peptide within the medial articular capsule. The fibres were predominantly located in the superficial layer of the capsule. They formed a dense innervation pattern, mainly accompanying blood vessels. Electron microscopy showed that most fibres were in close proximity to small arteries. The highest innervation density was found in parts of the capsule that were located over the epicondyle of the femur with 21 +/- 12 fibres per mm2 (mean +/- SD). In the tissue over the joint cleft this density was lower, with 11 +/- 6 fibres per mm2. In conclusion, the high innervation density of the knee joint capsule by nerve fibres containing calcitonin gene-related peptide supports the hypothesis of an important regulatory function of this peptide in normal tissue.

Animals↗
PubMed1997

Anatomy and histology of joint innervation.

Joint diseases are mostly accompanied with hyperalgesia and/or pain in the joint region. As about 60% of the knee joint innervation may be classified as nociceptive, the structure of joint afferents became a matter of interest. The function of a sensory nerve fibre may be defined by its location, its morphology and its biochemical composition. This review is therefore focused on the composition of articular nerves, the morphology of sensory endings, their content of mitochondria and neuropeptides, and the plasticity of neuropeptide synthesis during joint inflammation. As the knee joint of the cat belongs to the best examined articular tissue concerning the sensory function of its afferent innervation, most of the morphological data have also been determined in this model. The histological studies revealed a dense afferent innervation of all parts of joints. Most of the afferents terminate in non-corpuscular endings. Although the ultrastructure of these endings has been examined in great detail, up to now no morphological and biochemical details can be used to discriminate between mechanosensitive and nociceptive afferents. Only the differences in the content of mitochondria in single nerve fibres may be related to different sensory functions. The content of different neuropeptides in joint afferents highlights an additional efferent function of a proportion of joint afferents. The dense innervation with peptidergic joint afferents indicates an important, but up to now mainly unknown role of neuropeptides in a normal joint. Pathological conditions like joint inflammations lead to an enhanced release of neuropeptides into the joint tissue and a specific upregulation of neuropeptide biosynthesis in articular afferents.

Cartilage, Articular↗
PubMed1996

Bradykinin receptors in cultured rat dorsal root ganglion cells: influence of length of time in culture.

The endogenous nonapeptide bradykinin is a powerful substance which activates nociceptors, resulting in the sensation of pain in man. We used a newly developed non-radioactive method to detect bradykinin binding sites in isolated dorsal root ganglion cells with gold-labelled bradykinin. In a subpopulation of cells, gold-labelled bradykinin was bound in different quantities. The proportion of somata with bradykinin binding markedly depended on the length of time in culture. After 0.75 days, bradykinin was bound to 43% of somata. This proportion increased to 85% after 1.75 days and then decreased to 27% after 5.75 days. Bradykinin was bound to cells of all sizes, ranging from 40 to 2000 microns2 with a maximum of 200-300 microns2. In some cells, binding was also seen along the processes. No correlation was found between the soma size and the density of bradykinin binding. Blocking the bradykinin binding at the B1 receptor with (Des-Arg10)-Lys-bradykinin and at the B2 receptor with D-Arg(Hyp3-Thi5.8-D-Phe7)-bradykinin, respectively, revealed that in 0.75-day-old cultures no or only a very small amount of B1 receptors are present. In 1.75-day-old cultures, the marked increase in the proportion of cells with positive bradykinin binding is due to a de novo expression of the B1 receptor subtype and an up-regulation of the B2 receptor subtype. The selective or combined addition of specific B1 and B2 receptor ligands revealed that both receptor subtypes are co-localized. These data show that cultured sensory neurons express not only B2, but during a short period of time in culture also B1 receptors. The data allow us to hypothesize that a transient increase in bradykinin receptor expression might be caused by cell injury due to disruption of the axon. The injury-induced up-regulation of the receptor in vivo could cause physiological reactions.

Animals↗
PubMed1995

The effect of a unilateral inflammation at the rat's ankle joint on the expression of preprotachykinin-A mRNA and preprosomatostatin mRNA in dorsal root ganglion cells--a study using non-radioactive in situ hybridization.

In rats with an acute (2 days) and chronic (20 days) unilateral ankle joint inflammation (induced by Freund's complete adjuvant), the proportion of dorsal root ganglion cells containing preprotachykinin-A mRNA or preprosomatostatin mRNA was determined using non-radioactive in situ hybridization. At the acute stage of inflammation, the proportion of neurons containing preprotachykinin-A mRNA was similar to that in control rats. At the chronic stage, the proportion of neurons expressing preprotachykinin-A mRNA was significantly higher on the inflamed side than on the contralateral side. The proportion of dorsal root ganglion cells containing preprosomatostatin mRNA did not change. These data suggest that inflammation influences the synthesis of substance P but not of somatostatin in afferent neurons.

Acute Disease↗
PubMed1995

Somatostatin-like immunoreactivity in primary afferents of the medial articular nerve and colocalization with substance P in the cat.

The proportion of somatostatin-containing dorsal root ganglion cells innervating the knee joint of the cat via the medial articular nerve was determined by using retrograde labeling with fast blue and immunohistochemistry. Immunoreactivity was found in 8.6% of labeled cell bodies. In colchicine-treated ganglia, the proportion increased to 16.8%. Only small and intermediate-sized perikarya showed somatostatin-like immunoreactivity, indicating that this neuropeptide is synthesized predominantly in primary afferent units with unmyelinated sensory axons but may also be present in primary afferents with thinly myelinated sensory fibers. Colchicine treatment had no influence on the cell size distribution. Colocalization of somatostatin with substance P was determined by comparing the proportions of immunopositive dorsal root ganglion cells after incubation with antibodies against substance P or somatostatin or with a mixture of both. Substance P-like immunoreactivity was found in 18.1% (untreated ganglia) and 19.6% (colchicine treated ganglia) of the labeled neurons. After incubation with a mixed antibody solution, 18.2% of joint afferents in untreated and 19.9% of the cells in colchicine-treated ganglia were immunopositive. Comparing this result with the results obtained using somatostatin and substance P antibodies alone, one can calculate that both neuropeptides are colocalized in about 17% of the cat's knee joint afferents. About 3% of the neurons contain only substance P, whereas almost none of the neurons contain only somatostatin. Based on this fact, one can assume that both neuropeptides are coreleased in peripheral tissue as well as in the central nervous system.

Afferent Pathways↗
PubMed1995

Proportion of NADPH-diaphorase reactive sympathetic efferents innervating the normal and inflamed knee joint of the cat.

To determine whether postganglionic sympathetic efferents may be a source of nitric oxide in normal and inflamed knee joints, the distribution of NADPH-diaphorase was studied in retrogradely labelled neurons of the paravertebral sympathetic ganglia in the cat. In these ganglia, a strong NADPH-diaphorase staining reaction was found in 3-7% (segments L2-L5) and 14-17% (segments L6 and L7) of the neurons. Only 2.5 +/- 1.2% (mean +/- SD, 4 normal joints), of 2207 labelled neurons showed a positive staining reaction. During a unilateral inflammation (32 h), this proportion slightly increased to 5.0 +/- 1.2% (unaffected joints of 4 animals, 2510 labelled perikarya) and 5.4 +/- 2.8% (inflamed joints of 4 animals, 2578 labelled perikarya). However, these increases only attained statistical significance between the values obtained from control animals and from the normal sides of animals with a monoarthritis. Thus, the data indicate that in the sympathetic innervation of the normal and inflamed knee in the cat, the release of nitric oxide has little or no importance.

Amino Acid Oxidoreductases↗
PubMed1995

Fine sensory innervation of the knee joint capsule by group III and group IV nerve fibers in the cat.

Afferent group III and IV nerve fibers of the knee joint markedly differ in their responsiveness to mechanical stimulation, which may be reflected in the structure and location of their terminals. Therefore, in sympathectomized cats, the fine afferent innervation of the knee joint capsule was studied via ultrastructural three-dimensional reconstructions over distances of up to 300 microns. Small peripheral nerves and "free" (noncorpuscular) sensory nerve endings were found in a superficial layer of the outer fibrous part of the capsule, in the patellar retinaculum, and in the outer and inner surface layers of the medial collateral and patellar ligaments. Group III nerve fibers showed a proximal myelinated portion inside the nerve, an intermediate portion that lacks a myelin sheath and is only surrounded by perineurium, and a distal portion outside of the perineurium that forms the sensory ending proper. Group IV fibers showed only two distinct portions, an intraperineurial (proximal) and an extraperineurial (distal) portion without any further morphological differences. Outside of the perineurium, a network formed by Schwann cells ("Schwann cell reticulum") provides a pathway for the distal portion of the sensory axons. No distinct subgroups of the sensory terminal fibers could be defined according to the configuration of the Schwann cells and the nerve fiber terminals. Sensory terminals were located adjacent to different structures such as venous and lymphatic vessels, fat cells, and collagenous fibers. Distinct parts of the same terminal nerve fiber were found in close contact to a vessel wall; others were surrounded by dense collagenous tissue. Close to sensory endings, mast cells and mast cell-like cells were frequently found, indicating a functional relationship.

Animals↗
PubMed1995

A simple method for a specific retrograde labelling of dorsal root and sympathetic ganglion cells innervating the knee joint of the cat.

To examine the biochemical characteristics of primary afferents and sympathetic efferents innervating the normal and inflamed knee joint of the cat, the corresponding perikarya have to be labelled retrogradely. In the present study we used an injection of the fluorescent dye Fast Blue through the intact skin and the patellar ligament into the centre of the knee joint cavity and examined the efficiency of this kind of labelling. The segmental distributions of labelled cell profiles in cryostat sections of dorsal root ganglia (DRG) and paravertebral ganglia of the sympathetic trunk agreed with data obtained previously after applying the dye at the transected end of the two main articular nerves. The soma size distribution of labelled DRG cells was identical to previous results indicating that the perikarya labelled in the present study may be a representative proportion of the knee joint innervation. Axotomy of the medial and posterior articular nerves reduced the number of labelled cell profiles of about 85%. Taken together, these data indicate that an injection of Fast Blue into the knee joint cavity of the cat revealed a highly specific and efficient retrograde labelling of the neuronal cell bodies innervating the knee joint.

Amidines↗
PubMed1995

Non-radioactive localization of substance P binding sites in rat brain and spinal cord using peptides labeled with 1.4-nm gold particles.

We present here a new method for non-radioactive labeling of substance P (SP) to demonstrate the distribution of its binding sites in histological sections. The peptide was labeled at the primary amino group with a 1.4-nm gold particle. In Western blots of membrane fractions of rat spinal cord, specific binding occurred at 38 and 58 KD. This binding was competitively suppressed by adding the native peptide. In addition, the SP-gold conjugate was able to displace the corresponding 125I-labeled peptide from binding proteins. In histological sections, binding sites could be shown in various parts of rat brain and spinal cord. The distribution patterns were comparable to those found in studies using autoradiographic methods. Adding the native peptide or a neurokinin 1 receptor agonist markedly reduced labeling of the tissue, whereas only a slight reduction was obtained after adding neurokinin A. Therefore, SP could be specifically labeled with a 1.4-nm gold particle to demonstrate its binding sites. This new method combines the advantages of receptor-ligand affinity binding with a non-radioactive detection system. It can be used for labeling peptides in general and therefore offers an alternative or addition to other methods used in study of the distribution of membrane receptors.

Animals↗
PubMed1994

NADPH-diaphorase reactivity in articular afferents of a normal and inflamed knee joint in the cat.

The distribution of NADPH-diaphorase was studied in retrogradely labelled dorsal root ganglion cells innervating the knee joint of the cat. A strong staining reaction was found in 7.5 +/- 1.9% (mean +/- S.D. of 9 normal joints and 6393 labelled perikarya) of the articular afferents. An acute inflammation (32 h) significantly increased this proportion to 10.9 +/- 2.2% (mean +/- S.D. of 5 inflamed joints and 3933 labelled perikarya). The diameter distribution of the somata with a positive NADPH-diaphorase reaction ranged from 18 to 46 microns with a maximum at 24-28 microns. These data indicate that a small proportion of knee joint primary afferents may be able to release nitric oxide playing a role in synaptic transmission and in regulatory functions within the peripheral tissue under normal and pathophysiological conditions.

Amidines↗
PubMed1994

Combination of alkaline phosphatase in situ hybridization with immunohistochemistry: colocalization of calretinin-mRNA with calbindin and tyrosine hydroxylase immunoreactivity in rat substantia nigra neurons.

We describe a method to combine non-radioactive in situ hybridization using alkaline phosphatase (AP) labelled oligonucleotide-probes with immunohistochemistry on the same thin paraffin section. The simultaneous detection of calretinin-mRNA and calbindin- or tyrosine hydroxylase-like immunoreactivity in neurons of rat substantia nigra, pars compacta, was used as a test system to develop the method. Brains were fixed by perfusion with 4% paraformaldehyde and embedded in paraffin. Five-microns-thick sections were processed for non-radioactive in situ hybridization with a 33-base alkaline phosphatase conjugated synthetic oligonucleotide complementary to calretinin mRNA. After hybridization and colour reaction to visualize calretinin mRNA, sections were incubated with antibodies against calbindin D28K or tyrosine hydroxylase. Immunoreaction was visualized using the avidin-biotin-complex-technique and diaminobenzidine. As the colour of both reaction products differ markedly, the distribution of calretinin mRNA-containing neurons (purple-blue, alkaline phosphatase product) and calbindin/tyrosine hydroxylase immunopositive cells (brown peroxidase product) could be differentiated easily on the same section. Calbindin- and tyrosine hydroxylase-like immunoreactivity was found in the majority of calretinin mRNA-containing cells within the substantia nigra, pars compacta, indicating that in this nucleus a proportion of the dopaminergic neurons contain both calcium binding proteins calbindin and calretinin. In conclusion, non-radioactive in situ hybridization using alkaline phosphatase labelled oligonucleotide probes can be readily combined with immunohistochemistry.

Alkaline Phosphatase↗
PubMed1994

Mitochondria in fine afferent nerve fibres of the knee joint in the cat: a quantitative electron-microscopical examination.

The distribution of mitochondria, their content and concentration (expressed as the ratio of the mean volume of mitochondria and the surface of the sensory axon) were determined in group-III and -IV nerve fibres innervating the knee joint capsule in the cat. Mitochondria mainly accumulated in axonal swellings ("beads") and end bulbs of the terminal branches. Between single nerve fibres, marked differences in the content and the concentration of mitochondria were obtained in proximal portions (inside of the perineurium) and in distal portions (unmyelinated sensory endings). In group-III nerve fibres, the mitochondrial concentration ranged from 0.005 to 0.030 microns 3/microns 2 (proximal portion) and from 0.016 to 0.080 microns 3/microns 2 (distal portion). In unmyelinated group-IV nerve fibres, the values also showed a broad variation ranging from 0.001 to 0.011 microns 3/microns 2 (proximal portion) and from 0.003 to 0.019 microns 3/microns 2 (distal portion). The wide range of mitochondrial concentrations may reflect different energy consumption during receptive processes: nerve fibres with a low mechanical threshold and a high probability of excitatory events may be rich in mitochondria, whereas fibres with a high mechanical threshold and a low probability of excitatory events may be poor in mitochondria.

Animals↗
PubMed1993

Distribution of calretinin mRNA in rat spinal cord and dorsal root ganglia: a study using non-radioactive in situ hybridization histochemistry.

Using non-radioactive in situ hybridization calretinin mRNA was detected in numerous small neurons within lamina II and IV of the dorsal horn. Many labelled cells are distributed over the whole ventral horn; however, no motoneurons contained the mRNA. In dorsal root ganglia 4.9 +/- 1.7% (mean +/- S.D., n = 5 animals) of the primary afferent neurons contained calretinin mRNA. Labelled cells were of intermediate and large size with diameters ranging from 36 to 68 microns indicating that calretinin is synthesized in neurons with myelinated afferent fibers and presumably a corpuscular ending.

Animals↗
PubMed1992

Neurokinin A-like immunoreactivity in articular afferents of the cat.

Dorsal root ganglion cells with axons innervating the cat's knee joint via the medial articular nerve were retrogradely labelled with Fast blue. Neurokinin A-like immunoreactivity was found in 4.5 +/- 1.1% (mean +/- S.D. of 5 nerves and 695 cells) of the articular afferents. Colchicine treatment of the ganglia increased the percentage of immunopositive cells to 8.5 +/- 0.7% (mean +/- S.D. of 6 nerves and 554 cells) after 3-22 h. The diameter distribution of the immunopositive somata ranged from 20 to 50 microns with a maximum at 26-30 microns. Comparing the proportions of neurokinin A-immunopositive cells with those of substance P, it can be calculated on the basis of mRNA encoding that neurokinin A is synthetized in about half of the substance P-containing primary articular afferents.

Afferent Pathways↗
PubMed1991

Receptor-mediated endocytosis of albumin in cultured opossum kidney cells: a model for proximal tubular protein reabsorption.

The accumulation of fluorescein (FITC)-labelled bovine albumin was measured against the extracellular-fluid-phase marker FITC-inulin within confluent monolayers of the opossum kidney cell line OK. Fluorescence and electron microscopic pictures show that FITC-albumin is taken up by endocytosis and appears in a vesicular intracellular distribution. The uptake of FITC-albumin was quantified by measuring the cell-adherent fluorescence fluorimetrically. FITC-albumin uptake shows a time- and concentration-dependent saturation kinetics in contrast to the non-saturable FITC-inulin uptake, and exceeds the latter more than tenfold at low concentrations. Half-maximum saturation occurs at 20-30 mg/l. Initial FITC-albumin uptake/mg protein is stimulated by cell maturation, being six-to sevenfold higher in the confluent than in the subconfluent state, while FITC-inulin uptake is unchanged. Both an elevation of ambient osmolality to 600-750 mOsm/kg and disruption of the cytoskeleton by cytochalasin B (0.1 mmol/l) reduce initial FITC-albumin uptake by 50%-60% in a non-additive fashion. Albumin endocytosis is reduced both in acidic (pH 5.4) and alkaline (pH 8.4) medium, but does not depend on extracellular sodium, calcium or chloride. High concentrations of fetal calf serum or unlabelled albumin reduce FITC-albumin endocytosis dose-dependently. The present study is the first to investigate both the protein uptake and the fluid-phase endocytosis in a cultured proximal tubular cell line, using these cells as a model system-for proximal tubular protein reabsorption.

Albumins↗
PubMed1991

Substance P- and calcitonin gene-related peptide immunoreactivity in primary afferent neurons of the cat's knee joint.

The distribution of substance P and calcitonin gene-related peptide was determined in primary afferent neurons of the medial and posterior articular nerve of the cat's knee joint. Perikarya of articular afferents were visualized by retrograde labelling with the fluorescent dye Fast Blue which was applied at the transected end of the peripheral nerves. Substance P was found in about 17% of labelled medial articular afferents and in about 16% of labelled posterior articular afferents, respectively, whereas calcitonin gene-related peptide was present in about 35 and 32% of the medial and posterior articular nerve cells, respectively. Taking into account that these neuropeptides are known to be co-localized, probably not more than one-third of the joint afferents contain substance P and/or calcitonin gene-related peptide. Quantification of cell diameters revealed that substance P was found only in small- or intermediate-sized perikarya (less than 50 microns) indicating that this peptide is predominantly found in unmyelinated neurons. Calcitonin gene-related peptide was present mainly in small- and intermediate- but also in some large-sized neurons (greater than 50 microns) providing evidence that this peptide is found in unmyelinated and to a lesser extent in myelinated neurons. This is consistent with previous studies that show that substance P and calcitonin gene-related peptide are present primarily in unmyelinated and thinly myelinated primary afferents. When the portion of substance P-positive neurons of the medial articular nerve is compared to the number of articular afferents displaying a nociceptive function as determined in earlier electrophysiological studies, it can be calculated that at most 30% of the nociceptive-specific articular afferents contain this neuropeptide.(ABSTRACT TRUNCATED AT 250 WORDS)

Amidines↗
PubMed1991

Nociception and pain.

In recent years, progress in the treatment of pain has been strongly influenced by new insights into the mechanisms underlying pain and nociception. The following article briefly reviews some recent reports which make a significant contribution to our knowledge of the structure and function of nocisensors, the neuropeptides involved in the nociceptive system, and the spinal and supraspinal central mechanisms of nociception.

Afferent Pathways↗