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Biomedical subjects

A D Short

Publications and source records attributed to A D Short.

At least 37 records · Page 2Linked to original sources
PubMed1991

Persistent intracellular calcium pool depletion by thapsigargin and its influence on cell growth.

The intracellular Ca2+ pump inhibitor, thapsigargin, added to DDT1MF-2 smooth muscle cells in culture, irreversibly inhibited accumulation of Ca2+ within cells, permanently emptied the inositol 1,4,5-trisphosphate (InsP3)-sensitive Ca2+ pool, and simultaneously induced profound alteration of cell growth. After only a brief (30-min) treatment of cultured cells with 3 microM thapsigargin followed by extensive washing, the total releasable InsP3-sensitive Ca2+ pool remained entirely empty, even after 7 days of culture without thapsigargin. After thapsigargin treatment, cells retained viability, usual morphology, and normal mitochondrial function. Despite the otherwise normal appearance and function of thapsigargin-treated cells, cell division was completely blocked by thapsigargin. DNA synthesis was completely inhibited when thapsigargin was added immediately after passaging, but was suppressed only slowly (4-6 h) when added to rapidly synthesizing cells (24 h after passaging). Protein synthesis was reduced by approximately 70% in thapsigargin-treated cells. The sensitivity of thapsigargin-mediated inhibition of cell division, DNA synthesis, protein synthesis, and Ca(2+)-pumping activity were all similar with the EC50 values for thapsigargin in each case being close to 10 nM. Upon application to DDT1MF-2 cells, thapsigargin transiently increased resting cytosolic Ca2+ (0.15 microM) to a peak of 0.3 microM within 50 s; thereafter, free Ca2+ declined to 0.2 microM by 150 s and continued to slowly decline toward resting levels. Cells treated with thapsigargin for 1-72 h in culture displayed normal resting cytosolic Ca2+ levels. However, application of thapsigargin or epinephrine to such cells resulted in no change in the intracellular Ca2+, indicating that the internal Ca2+ pool remained completely empty. These results suggest that emptying of Ca2+ from intracellular thapsigargin-sensitive Ca(2+)-pumping pools induces profound alteration of cell proliferation.

Animals↗
PubMed1991

Direct observations of synapses between GABA-immunoreactive boutons and identified spinocervical tract neurons in the cat's spinal cord.

Three spinocervical tract neurons in adult cats were physiologically characterized and intracellularly labelled with horseradish peroxidase. The neurons were reconstructed and examined with the light microscope and were prepared for postembedding immunochemical analysis by using an antiserum which specifically recognizes GABA in glutaraldehyde-fixed tissue. Semithin sections were tested and examined with the light microscope. Somata, proximal, and distal dendrites of all three cells were associated with numerous punctate GABA-immunoreactive structures. Immunoreactive perikarya of small neurons in the vicinity of spinocervical tract cells were also observed. Ultrastructural analysis, with the immunogold technique, revealed that somata and proximal dendrites of all three neurons received synaptic contacts (about 37% of total synapses) from GABA-immunoreactive boutons and that distal dendrites were also associated with substantial numbers of immunoreactive structures (about 27% of synapses). Immunoreactive boutons were small (about 1 micron in diameter), contained irregularly shaped agranular vesicles, and formed symmetrical synaptic junctions with identified neurons. An additional group of immunoreactive boutons was observed to be associated with one of the cells only; these contained many large dense-core vesicles in addition to small agranular vesicles. Boutons containing round agranular vesicles and flattened agranular vesicles were not observed to be immunoreactive. The evidence supports the idea that much of the postsynaptic inhibition observed in spinocervical tract neurons is mediated by GABA and that even the most distal dendrites of these neurons receive inhibitory inputs.

Animals↗
PubMed1991

The effect of retinoic acid on parathyroid hormone- and parathyroid hormone-related peptide-induced intracellular calcium in a rat osteosarcoma cell line, UMR106.

We have examined the effects of parathyroid hormone (PTH) and PTH-related peptide (PTH-rP) on intracellular calcium (Ca2+i) in a rat osteogenic sarcoma cell line, UMR106. Synthetic bovine (b)PTH(1-34) caused a small inconsistent rise in Ca2+i in UMR106 cells, whilst cells pretreated with retinoic acid (RA, 1 mumol/l) for 18 h exhibited reproducible, significant and dose-dependent increases in Ca2+i levels in response to bPTH. The effect of RA on PTH-induced changes in Ca2+i were dependent upon both dose and time. Purified human (h)PTH-rP(1-34) increased Ca2+i in the absence of RA in the same cells. However, RA increased the magnitude of PTH-rP-stimulated changes in Ca2+i without affecting the concentration required for a maximal response. RA also prolonged the delay before the Ca2+i response was observed. Maximal responses to PTH-rP were greater in magnitude than those to PTH. These changes appeared not to be due to cyclic AMP (cAMP), since neither dibutyryl cAMP (1 mmol/l) nor forskolin (15 mumol/l) affected Ca2+i. PTH- and PTH-rP-mediated Ca2+i transients were not completely abolished by the absence of extracellular calcium, and both peptides increased basal levels of inositol trisphosphate. PTH and PTH-rP were subject to mutual desensitization, but were not desensitized by prostaglandin E2. PTH(7-34) antagonized PTH- but not PTH-rP-mediated Ca2+i transients. We conclude that there may be some important differences in the mechanism of action of PTH and PTH-rP.

Animals↗
PubMed1990

Direct observations of synapses between GABA-immunoreactive boutons and muscle afferent terminals in lamina VI of the cat's spinal cord.

Single group Ia muscle afferent fibres in the lumbar spinal cord of the cat were impaled with microelectrodes and labelled with horseradish peroxidase. Two collateral axons were prepared for combined light and electron microscopy. Arbors selected from lamina VI were processed by the postembedding immunogold technique with antiserum which specifically recognizes GABA in glutaraldehyde-fixed tissue. Twelve Ia boutons were examined through series of thin sections with the electron microscope and all of them were associated with presynaptic axon terminals which were positively labelled for GABA. Some Ia boutons received synaptic contacts from several GABAergic terminals. The present study establishes that a GABA-like substance is present in axon terminals presynaptic to Ia afferent boutons in lamina VI of the spinal cord. This evidence provides a morphological basis for presynaptic inhibition of Ia afferent input into lamina VI.

Animals↗
PubMed1990

Interleukin-1 stimulates diglyceride accumulation in the absence of protein kinase C activation.

Despite advances in the knowledge of the intracellular signalling in response to extracellular messengers, the mechanism of action of interleukin-1 (IL-1) has remained an enigma. In the present study, we have employed human dermal fibroblasts (Detroit 532 cells) to investigate IL-1 beta-induced changes in intracellular signals. Both recombinant human IL-1 beta and a native preparation purified from human placental tissue were employed. Cyclic AMP levels in cell monolayers were unaltered by IL-1 beta. Also, IL-1 beta did not influence significantly the levels of phosphatidylinositol, phosphatidylinositol 4-monophosphate, and phosphatidylinositol 4,5-bisphosphate in the membrane, nor the water-soluble inositol phosphates, inositol monophosphate, inositol bisphosphate and inositol trisphosphate, in cells prelabelled with myo-[3H]inositol. In addition, intracellular calcium as measured by Quin2 was unaffected by interleukin-1. However, in cells labelled with [3H]glycerol or [3H]arachidonic acid, IL-1 beta caused an immediate rise in diglyceride (DG) accumulation. As the effects of IL-1 beta have been reported to be mimicked by tumour-promoting phorbol esters, this rise in DG suggested the involvement of protein kinase C (PKC). However, repeated experiments failed to reveal any acute effect of IL-1 beta on the activity of this enzyme. Furthermore, IL-1 beta did not cause the translocation of PKC between the membrane and the cytosol as has been found in response to other extracellular signals. Rather, IL-1 beta appeared to increase the synthesis of PKC in both membrane and cytosol preparations, an effect which could be prevented by coincubation with cycloheximide. These findings suggest that the diglyceride formed in response to IL-1 beta does not activate protein kinase C.

Calcium↗
PubMed1990

Interactive regulation of signalling pathways in bone cells: possible modulation of PGE2-stimulated adenylyl cyclase activity by protein kinase C.

We have reported previously that tumour-promoting phorbol esters modulate both basal and vasoactive intestinal polypeptide (VIP)-stimulated adenylyl cyclase activity in GH3 (an established pituitary cell line). Here, we probe the receptor and cell specificity of this response. Experiments were performed in the presence of isobutylmethylxanthine. Unlike the response in GH3 cells, the tumour-promoting phorbol ester (tetradecanoyl phorbol acetate (TPA] did not affect either basal adenylyl cyclase activity nor VIP-stimulated activity in the rat osteosarcoma subclones UMR 106-01 and UMR 106-06. In addition, the cyclase responses to parathyroid hormone (PTH), and, in the case of UMR 106-06, to calcitonin were unaffected by tumour-promoting phorbol ester. However, prostaglandin E2-stimulated cyclase activity in both of these subclones was attenuated in a dose-dependent manner.

Adenylyl Cyclases↗
PubMed1990

Central boutons of glomeruli in the spinal cord of the cat are enriched with L-glutamate-like immunoreactivity.

Several lines of evidence indicate that L-glutamate may be a neurotransmitter of fine myelinated and unmyelinated primary afferent fibres in the spinal cord. The aim of the present study was to determine if L-glutamate was enriched in the terminals of these fibres. We performed the post-embedding immunogold technique on sections taken from the superficial regions of the lumbar cord in two cats. An antiserum, raised against protein-conjugated L-glutamate, was employed. Several tests on tissue and on a model system indicated that the antiserum recognized a glutaraldehyde-fixed L-glutamate-like substance. Terminals of fine afferent fibres were identified in the substantia gelatinosa as central boutons of synaptic glomeruli. Central boutons were examined through serial sections following immunogold reactions and were found to be heavily labelled with gold particles in consecutive sections. Quantitative analysis indicated that central boutons were more than two and a half times as densely labelled with gold particles than the tissue average. It was concluded that this represents a genuine enrichment of L-glutamate in these structures. Comparisons were made between L-glutamate-immunoreactive properties of central terminals and immunoreactivity for GABA, aspartate and glutamine. Statistical analysis revealed that central boutons in sections incubated in GABA antiserum and glutamine antiserum were associated with significantly lower densities of gold particle labelling than the average for the same tissue. Particle densities of central boutons in sections incubated in aspartate antiserum were not significantly different from average tissue densities. It was concluded that central boutons were not enriched with these three amino acids. Central boutons of synaptic glomeruli were classified into three groups on morphological criteria: (1) dense sinusoidal boutons; (2) large dense-core vesicle-containing boutons; and (3) regular synaptic vesicle-containing boutons. Quantitative analysis revealed that all of these groups were enriched in glutamate immunoreactivity, however, there were differences between the groups; large dense-core vesicle-containing boutons were associated with significantly lower densities of particles than regular synaptic vesicle-containing and dense sinusoidal terminals. The evidence indicates that central boutons, which most probably originate from fine myelinated and unmyelinated primary afferent fibres, are enriched with L-glutamate which may serve as a neurotransmitter in such fibres.

Animals↗
PubMed1990

Afferent inhibition and facilitation of transmission through the spinocervical tract in the anaesthetized cat.

1. Extracellular microelectrode recordings were made from single spinocervical tract (SCT) neurones in the lumbosacral spinal cord of cats anaesthetized with chloralose and paralysed with gallamine triethiodide. 2. Pairs of air-jet stimuli, 60 ms in duration, were used to investigate in-field afferent inhibition in SCT cells. One jet was used to condition the responses to another jet located at a different position within the excitatory receptive field and occurring at times from 100 to 1800 ms later. Fifteen neurones were tested and significant in-field inhibition was observed in all of them. 3. The in-field afferent inhibition was organized spatially in the sense that inhibition was generally strongest when conditioning and testing stimuli were close together and became weaker as they were moved apart. There was also a weak effect due to the strength of the conditioning response; when conditioning produced a strong response, from near the most excitable part of the receptive field, there was often a weak reduction in the test response from distant sites. The inhibitory areas defined in these experiments were generally less than 100 mm in length in units with excitatory receptive fields much longer than this. 4. The in-field afferent inhibition had a time course that lasted from 300 to about 1000 ms. 5. Afferent inhibition was also evoked by applying either air-jet stimuli to hairy skin outside, but close to, the excitatory receptive field or by applying a vibratory stimulus from a piezoelectric transducer (200 Hz) to glabrous skin of the toe pads or the central foot pad. These conditioning stimuli had durations of 20 or 60 ms. For convenience we call this inhibition 'out-of-field' afferent inhibition. 6. Out-of-field afferent inhibition was evoked from both glabrous and hairy skin areas outside the excitatory receptive field. It was common in neurones with receptive fields on the toes and of twenty-eight such neurones tested it was observed in twenty-four. This inhibition had a short latency (usually about 10 ms or less but occasionally up to 30 ms) and lasted for about the duration of the test stimulus (30 or 80 ms when the test stimulus was 20 or 60 ms respectively). It was often followed by a further period of inhibition, with a latency of between 50 and 100 ms and lasting for 60 up to 130 ms. 7. In thirteen SCT neurones more complex effects were seen.(ABSTRACT TRUNCATED AT 400 WORDS)

Action Potentials↗
PubMed1989

Spatial spread of in-field afferent inhibition in the cat's spinocervical tract.

1. Extracellular microelectrode recordings were made from twenty-three spinocervical tract (SCT) cells in the lumbar spinal cord of cats anaesthetized with chloralose and paralysed with gallamine triethiodide. Excitation and inhibition of the cells were elicited by applying small brief (4 mN, 60 ms) localized jets of air to the clipped hair in and around the receptive fields. 2. Receptive field extents ranged from 40 to 180 mm. Excitation occurred in the period 30-130 ms after the start of the stimulus, and in-field afferent inhibition from 130 ms up to 700 ms or more. The inhibition was manifest as a reduction in background discharge and as a reduction in responsiveness to a test stimulus which followed a conditioning stimulus. 3. When the conditioning stimulus was spatially separated from the test stimulus, the degrees of in-field afferent inhibition depended on the spatial separation, even when both were within the excitatory receptive field. The spatial spread of in-field afferent inhibition was limited to 100 mm or less. 4. In two units only, afferent inhibition was produced from a narrow strip just outside the excitatory receptive field. In the other units, it could only be produced from within the excitatory receptive field. 5. The results suggest that the inhibitory input to SCT cells is organized in subdomains no more than 100 mm across, which may correspond to the receptive fields of interneurones between the primary afferent fibres and the SCT cells.

Afferent Pathways↗
PubMed1989

Receptive fields and in-field afferent inhibition of neurones in the cat's lateral cervical nucleus.

1. Extracellular microelectrode recordings were made from projection neurones of the lateral cervical nucleus (LCN) in cats anaesthetized with chloralose and paralysed with gallamine triethiodide. 2. The receptive fields of eight-five units were analysed. Most units had excitatory receptive fields similar in size and shape to those of spinocervical tract (SCT) cells. A few (14%) had either very large fields or 'stocking-like' fields. The majority of the LCN neurones (fifty-five, 65%) were excited by hair movement and, in addition, by noxious mechanical stimulation within the skin area responding to hair movement. Twenty-five units (29%) were excited by hair movement alone. For seven of these twenty-five neurones, noxious mechanical stimulation within the excitatory receptive field produced inhibition of the background discharge. One unit was excited by noxious mechanical stimulation and for the remaining four units no receptive field could be found. In six units inhibitory receptive fields outside the excitatory field were found. 3. Air-jet stimuli were used to define the excitatory profiles of the units' receptive fields to hair movement. In general, receptive fields had single regions of greatest sensitivity usually at or near the centre of the field, where that was oval in shape, with the sensitivity declining towards the field's circumference. In some units with very large fields that included parts of one or two limbs and the trunk there could be more than one highly sensitive region. 4. Pairs of air-jet stimuli were used to investigate in-field afferent inhibition in LCN cells. One jet was used to condition the responses to another jet located at a different position within the excitatory receptive field and occurring 200 ms later. Sixteen units were tested and significant in-field inhibition was observed in all sixteen. 5. The in-field afferent inhibition was organized spatially in the sense that inhibition was generally strongest when the conditioning and testing stimuli were close together and became weaker as they were moved apart. The afferent inhibition was not simply a function of the response produced by the conditioning stimulus. Furthermore, increasing the strength of the stimuli did not in general lead to larger areas from which the inhibition could be produced. The inhibitory areas defined in these experiments were generally less than 120 mm in length in units with receptive fields much longer than 100 mm.(ABSTRACT TRUNCATED AT 400 WORDS)

Afferent Pathways↗
PubMed1978

Peritoneal mesothelioma in a male pseudohermaphrodite with asymmetrical gonadal differentiation.

A diffuse peritoneal mesothelioma occurring in a patient with male pseudohermaphroditism and asymmetrical gonadal differentiation (mixed gonadal dysgenesis) is described. Malignancies of mesodermal origin, usually derivatives of the urogenital ridge, appear to occur with increased frequency in male pseudohermaphrodites. This appears to be the first reported instance of a mesothelioma in a male pseudohermaphrodite with mixed gonadal dysgenesis. The clinical and pathologic features of this tumor, including the electron microscopic findings, are presented.

Disorders of Sex Development↗
PubMed1977

Inhibition of spinocervical tract discharges from localized areas of the sensorimotor cortex in the cat.

1. Intracortical microstimualtion (ICMS) was applied within the sensorimotor cortex of cats anaesthetized with chloralose. 2. The effects of the ICMS were examined on the number of impulses in spinocervical tract (SCT) cells (recorded extracellularly in the contralateral lumbosacral spinal cord) evoked by peripheral stimulation. 3. Inhibition of SCT discharges was produced by ICMS in two distinct regions of the sensorimotor cortex. 4. One inhibitory regions was in part of cytoarchitectonic area 4 gamma in the upper bank of the cruciate sulcus. It sometimes extended caudally into area 4 delta, medially into area 3 alpha and/or rostrally into the part of area 4 gamma on the caudal lip of the cruciate sulcus. 5. The other inhibitory region was in the medial part of the posterior sigmoid gyrus and included parts of areas 3 alpha, 3 beta, 1, 5 alpha and 5 beta. 6. Most inhibitory sites were in cortical layers III, V and VI. 7. No regions were found in which ICMS consistently caused facilitation of SCT discharges.

Animals↗
PubMed1970

Interaction of cortical evoked potentials in the rat.

1. Unitary and mass potentials were recorded with glass micropipettes at different depths in and around the primary somatosensory area of the cortex in rats anaesthetized with urethane; in addition, surface mass potentials were recorded with chlorided silver ball electrodes. Potentials were evoked by stimuli to the contralateral forepaw and the contralateral cortex. Observations were confined to potentials evoked within 20 msec of stimulation.2. If forepaw stimuli were applied at times when the cortex was not showing spontaneous activity, one component of the evoked response was a widespread depth negativity. This was accompanied by a smaller surface positivity which had the same time course. It could be recorded over the same area of cortex. Unitary activity was found with the same latent period and spatial distribution as the depth negativity. These components of the evoked mass and unitary responses to forepaw stimulation were absent if times of spontaneous cortical activity were chosen for delivering the stimuli.3. Contralateral cortical stimuli evoked mass activity similar to the component of the mass response to forepaw stimulation described in 2. Stimuli given simultaneously to the two sites elicited less unitary activity than the sum of the unitary activity evoked by the two stimuli separately. The mass activity exhibited the interactions which would be expected on the hypothesis that it is generated by the unitary activity. The time course of the interaction is described.4. The interaction was confined to the components of the evoked potentials which were only present if stimuli were applied when the cortex was quiescent. Stimuli applied when the cortex was active evoked mass and unitary potentials which showed no interaction.

Animals↗