Presynaptic and postsynaptic organelles of synapses formed in cultures of previously dissociated mouse spinal cord

Summary This study describes some of the ultrastructural features of presynaptic and postsynaptic organelles at synapses developed in cultures of previously dissociated mouse spinal cord cells. Particular attention was paid to the agranular reticulum which is well developed at many presynaptic and postsynaptic sites, either in the form of simple tubules or cisternae, or more complex networks and often closely associated with mitochondria. In addition, the disposition of microtubules at and close to synaptic specializations is described. These and other features of synaptic zones, such as granular vesicles in presynaptic sites, are discussed in relation to cultures developed on feeder layers and synapses in vivo, and in relations to possible degenerative and regenerative events in the cell cultures.     

Regional specificity of functional sensory connections in developing spinal cord cultures varies with the incidence of spontaneous bioelectric activity

Summary Cultured spinal cord explants in which little spontaneous bioelectric activity was present showed, when monitored using sensory ganglion-evoked monosynaptic action potentials, diffuse innervation by ingrowing afferent fibers at 3–4 weeks in vitro. In contrast, highly active cultures of the same age showed a strong tendency for functional sensory connections to be made within the dorsal half of the cord. Regional specificity was present in mature cultures (4–5 weeks in vitro), however, even when their spontaneous activity level was low. The results support earlier results using tetrodotoxin, and make it appear likely that centrally generated neuronal discharges can influence the topography of afferent terminals within the developing spinal cord.     

The vincamine derivative vindeburnol provides benefit in a mouse model of multiple sclerosis: effects on the Locus coeruleus

The endogenous neurotransmitter noradrenaline (NA) plays several roles in maintaining brain homeostasis, including exerting anti-inflammatory and neuroprotective effects. The primary source of NA in the CNS are tyrosine hydroxylase (TH)-positive neurons located in the Locus coeruleus (LC) which send projections throughout the brain and spinal cord. We recently demonstrated that dysregulation of the LC:Noradrenergic system occurs in experimental autoimmune encephalomyelitis as well as in MS patients, associated with damage occurring to LC neurons. Vindeburnol, a structural analog of the cerebral vasodilator vincamine, was previously reported to increase TH expression and activity in LC neurons. Female C57BL/6 mice were immunized with myelin oligodendrocyte glycoprotein (MOG)(35-55) peptide, and treated with vindeburnol at the first appearance of clinical signs. Clinical signs continued to increase for about 1 week, at which point mice in the vehicle group continued to worsen while vindeburnol-treated mice showed improvement. Pro-inflammatory cytokine production from splenic T cells was not reduced by vindeburnol suggesting primarily central actions of treatment. In the cerebellum, vindeburnol decreased astrocyte activation and reduced the number of demyelinated regions. Vindeburnol reduced astrocyte activation in the LC, reduced TH+ neuronal hypertrophy, increased expression of several genes involved in LC survival and maturation, and increased NA levels in the spinal cord. These results suggest that treatments with drugs such as vindeburnol which target LC survival or function could be of benefit in MS patients.     

依托咪酯对成年大鼠脊髓胶状质局部突触传递的作用

应用盲插全细胞膜片钳技术,在成年大鼠脊髓薄片上观察依托咪酯(etomidate,ET)对脊髓胶状质局部突触传递的影响。实验结果显示,在钳制电压为-70mV时,500μmol/L的ET对微小兴奋性突触后电流(mEPSC)的持续时间、频率和幅度都无明显的作用。在钳制电压为0mV时,50μmol/L的ET使GABA能微小抑制性突触后电流(mIPSC)的持续时间延长45.57±12.46%(P<0.05),但对其频率和幅度无影响。同样在钳制电压为0mV的情况下,50μmol/L的ET对甘氨酸能mIPSC的持续时间、频率及幅度均无作用。以上结果表明,在成年大鼠的脊髓胶状质,ET主要通过延长GABA能mIPSC的持续时间,即延长受体通道的开放时间发挥作用,ET对于兴奋性的突触传递没有直接的作用。     

Impact of acute and chronic stressor experiences on heart atrial and brain natriuretic peptides in response to a subsequent stressor

The impact of stressful events on processes related to cardiovascular functioning might vary with previous stressor experiences, just as such sensitization effects have been detected with respect to several neurochemical and hormonal processes. The present investigation assessed the impact of a psychosocial stressor on factors directly or indirectly related to cardiovascular functioning among CD-1 mice that had previously experienced an acute or chronic stressor regimen. These factors included plasma variations of atrial and brain natriuretic peptides (ANP and BNP, respectively), inflammatory cytokines in plasma, mRNA expression of natriuretic peptides and inflammatory cytokines in the ventricles, and norepinephrine (NA) levels and utilization within the locus coeruleus, a brain region implicated in cardiac functioning. A social stressor (exposure to a dominant mouse) increased NE levels and utilization within the locus coeruleus, plasma corticosterone, cytokine and ANP levels. Among mice initially exposed to an acute stressor (restraint), NE utilization, ventricular ANP mRNA expression, and plasma interleukin-6 (IL-6) concentrations were markedly increased by the subsequent social stressor. In chronically stressed mice some of the effects of the social stressor were dampened, including changes of plasma corticosterone, locus coeruleus NE utilization, as well as plasma and ventricular IL-6 mRNA expression. Conversely, plasma ANP was markedly enhanced by the combined stressor events as was ventricular BNP and IL-1β mRNA expression. It seems that stressors may profoundly influence (sensitize or desensitize) on factors that could influence cardiovascular functioning. It remains to be determined whether these actions would be translated as pathophysiological outcomes.     

Topical application of dynorphin A (1-17) antiserum attenuates trauma induced alterations in spinal cord evoked potentials,microvascular permeability disturbances,edema formation and cell injury: an experimental study in the rat using electrophysiological and morphological approaches

Summary.  Dynorphin is a neuropeptide that is present in high quantities in the dorsal horn of the spinal cord. The peptide is actively involved in pain processing pathways. However, its involvement in spinal cord injury is not well known. Alteration in dynorphin immunoreactivity occurs following a focal trauma to the rat spinal cord. Infusion of dynorphin into the intrathecal space of the cord results in ischemia, cell damage and abnormal motor function. Antibodies to dynorphin when injected into the intrathecal space of the spinal cord following trauma improve motor recovery, reduce edema and cell changes. However, influence of dynorphin on trauma induced alteration in spinal cord bioelectrical activity is still not known. Spinal cord evoked potentials (SCEP) are good indicator of spinal cord pathology following trauma. Therefore, in present investigation, influence of dynorphin antibodies on trauma induced changes in SCEP were examined in our rat model. In addition, spinal cord edema formation, microvascular permeability disturbances and cell injury were also investigated. Our results show that topical application of dynorphin antiserum (1 : 200) two min before injury markedly attenuated the SCEP changes immediately after injury. In the antiserum treated animals, a significant reduction in the microvascular permeability, edema formation and cell injury was observed in the traumatised spinal cord. These observations suggest that (i) dynorphin is involved in the altered bioelectrical activity of the spinal cord following trauma, (ii) the peptide actively participates in the pathophysiological processes of cell injury in the spinal cord trauma, and (iii) the dynorphin antiserum has potential therapeutic value for the treatment of spinal cord injuries. Received July 3, 2001 Accepted August 6, 2001 Published online July 31, 2002     

Between-day reliability of MyotonPRO for the non-invasive measurement of muscle material properties in the lower extremities of patients with a chronic spinal cord injury

Measuring the muscle properties of patients with spinal cord injuries (SCIs) is important to better understand their biomechanical features. In this study, we sought to evaluate the between-day reliability of MyotonPRO, a handheld device that can measure muscle mechanical properties, and assess whether it is reliable to measure muscle properties over time in patients with SCI. Thirteen men with complete SCIs (age 53.9 ± 6.3 years, height 171.0 ± 5.2 cm, weight 66.1 ± 5.8 kg), and injury levels ranging from L1 to T12, were enrolled. Oscillation frequency; logarithmic decrement; dynamic stiffness; mechanical stress relaxation time; and creep of the biceps femoris, medial and lateral gastrocnemius, rectus femoris, tibialis anterior, and Achilles tendon were measured on consecutive days using MyotonPRO. The intraclass coefficient for most muscles and the Achilles tendon ranged from 0.53 to 0.99 for all parameters. The percentage standard error of the measurement for many parameters in most muscles and the Achilles tendon was less than 10%. Bland-Altman analysis showed a high agreement for all mechanical properties. No significant differences were observed in any muscle or Achilles tendon properties between days (all p > 0.05). These results indicate that the MyotonPRO is reliable for between-day measurements of the mechanical properties of lower limb muscles and Achilles tendon in patients with SCI.     

Effect of maximal arm exercise on skin blood flux in the paralyzed lower limbs in persons with spinal cord injury

The purpose of the present study was to examine the effect of maximal arm exercise on the skin blood circulation of the paralyzed lower limbs in persons with spinal cord injury (PSCI). Eight male PSCI with complete lesions located between T3 and L1 performed graded maximal arm-cranking exercise (MACE) to exhaustion. The skin blood flux at the thigh (SBFT) and that at the calf (SBFC) were monitored using laser-Doppler flowmeter at rest and for 15 s immediately after the MACE. The subject's mean peak oxygen uptake and peak heart rate was 1.41 ± 0.22 1·min⁻¹ and 171.6 ± 19.2 beats·min⁻¹, respectively. No PSCI showed any increase in either SBFT or SBFC after the MACE, when compared with the values at rest. These results suggest that the blood circulation of the skin in the paralyzed lower limbs in PSCI is unaffected by the MACE.     

Ascending curbs of progressively higher height increases forward trunk flexion along with upper extremity mechanical and muscular demands in manual wheelchair users with a spinal cord injury

High upper extremity (U/E) demands are required when manual wheelchair users (MWUs) with spinal cord injury (SCI) ascend curbs; this may contribute to the risk of developing U/E musculoskeletal impairments. The aim of this study was to compare movement strategies (kinematics), mechanical loads (kinetics) and muscular demand (EMG) at the non-dominant U/E among 15 MWUs with SCI when ascending curbs of 4 cm (3 trials), 8 cm (3 trials) and 12 cm high (3 trials) from a starting line set 3 m before the curb. Biomechanical data was collected during three trials for each height. The curb ascent task was divided into three adjustment phases: caster pop, rear-wheel ascent and post-ascent. The greatest effort was generated by the shoulder flexors and internal rotators as well as the elbow flexors. A significant difference (p < 0.0167) between the curb heights was found for most outcome measures studied: movement excursion, net joint moments and muscular utilization ratio (MUR) of the main muscles increased with the higher curb heights, mainly around the shoulder joint. These results provide insight that aside from adhering to a highly structured training method for wheelchair curb ascent, rehabilitation professionals need to propose task-specific strength training programs based on the demands documented in this study and continue to advocate for physically accessible environments.     

Heat shock proteins as biomarkers for the rapid detection of brain and spinal cord ischemia: a review and comparison to other methods of detection in thoracic aneurysm repair

The heat shock proteins (HSPs) are members of highly conserved families of molecular chaperones that have multiple roles in vivo. We discuss the HSPs in general, and Hsp70 and Hsp27 in particular, and their rapid induction by severe stress in the context of tissue and organ expression in physiology and disease. We describe the current state of knowledge of the relationship and interactions between extra- and intracellular HSPs and describe mechanisms and significance of extracellular expression of HSPs. We focus on the role of the heat shock proteins as biomarkers of central nervous system (CNS) ischemia and other severe stressors and discuss recent and novel technologies for rapid measurement of proteins in vivo and ex vivo. The HSPs are compared to other proposed small molecule biomarkers for detection of CNS injury and to other methods of detecting brain and spinal cord ischemia in real time. While other biomarkers may be of use in prognosis and in design of appropriate therapies, none appears to be as rapid as the HSPs; therefore, no other measurement appears to be of use in the immediate detection of ongoing severe ischemia with the intention to immediately intervene to reduce the severity or risk of permanent damage.     

Simulation of the bis-(penicillamine) enkephalin in ammonium chloride solution: a comparison with sodium chloride

In order to quantify specific ion effects, a simulation study of bis(penicllamine) enkephalin, also known as DPDPE, has been performed in aqueous ammonium chloride solution and has been compared to a previous simulation of DPDPE in aqueous sodium chloride solution. Global thermodynamics have been calculated for a model system and the solution environment around DPDPE has been characterized. Associations of ions with DPDPE have been investigated. The observed differences between sodium chloride solution and ammonium chloride solution suggest that individual cations affect the solvation and peptide binding properties of a given anion.     

Effect of arm cranking exercise on skin blood flow of lower limb in people with injuries to the spinal cord

The purpose of this study was to examine whether arm cranking exercise induces changes in skin blood flow in the paralyzed lower limbs of people with injuries to the spinal cord (PISC). Ten PISC with lesions located between Th5 and L5 and six control subjects performed arm cranking exercise for 6 min at three intensities, 10, 30 and 50 W, at a room temperature of 25°C. Oxygen uptake (Vo₂) and heart rate (HR) were measured for the last 2 min of each exercise period. The skin blood flow at the anterior thigh (BF_sk,t) was continuously monitored using laser Doppler flowmetry for the whole 6-min period and for the first 10 min of recovery following exercise. During exercise, the PISC showed lower Vo₂ and greater HR than the control subjects. No increase in BF_sk,t was found in six of the PISC with lesions at or above Th12, irrespective of the exercise intensity. On the other hand, in PISC with lesions at L1 or below, BF_sk,t increased significantly (P < 0.05) with an increase in Vo₂ and HR, although the BF_sk,t at a given Vo₂ and HR was lower than that in the control subjects. These results would suggest that arm exercise can promote the blood circulation in the skin of the lower limbs if the injury level is below L1.     

Role of calcium ions in the stimulatory actions of luteinizing hormone in isolated ovarian cells: Studies with divalent-cation ionophores

The divalent-cation ionophores A23187 and ionomycin exert dose-dependent suppressive effects on the stimulatory actions of luteinizing hormone in ovarian cells $\text{in}\text{vitro}$. Micromolar concentrations of both A23187 and ionomycin can inhibit the production of progesterone and the stimulation of ornithine decarboxylase activity. Inhibitory concentrations of these ionophores deplete total cell content of calcium, and also seem to suppress protein synthesis.     

The influence of animal sex hormones on the induction of flowering in Arabidopsis thaliana: comparison with the effect of 24-epibrassinolide

The influence of selected steroids on the in vitro generative development of Arabidopsis thalianawas investigated. The activity of the animal steroids androsterone, androstenedione, progesterone, estrone, estriol, and 17-estradiol was compared to 24-epibrassinolide, a member of the regulatory family of brassinosteroids. A. thaliana plants were cultured in vitro in media containing these steroids. The stimulatory effect of the tested substances was evaluated by measurement of the percentage of generative plants versus vegetative plants in the experimental group. It was established that androstenedione, the main testosterone precursor, and androsterone, a typical male hormone, were more effective in stimulating flowering in A. thaliana than the female hormones, estrogens and progesterone. Androsterone at a concentration of 0.1 M increased the percentage of generative plants up to 96% (control 41%). Estrogens at the same concentration decreased the number of generative plants and 24-epibrassinolide did not stimulate A. thalianagenerative development.     

Characterization of a Novel Serotonin Receptor Subtype (5-HTls) in Rat CNS: Interaction with a GTP Binding Protein

Three pharmacologically distinct high-affinity [3H]serotonin ([3H]5-HT) binding sites were identified in spinal cord synaptosomes. [3H]5-HT competition studies using selective 5-HT1A receptor ligands indicated that approximately 25% of high-affinity synaptosomal [3H]5-HT binding was inhibited by 5-HT1A-selective compounds, an estimate consistent with [3H](+-)-8-hydroxy-2-(di-n-propylamino)tetralin ([3H]8-OH-DPAT) saturation experiments in which 5-HT1A receptors were directly labeled. [3H]5-HT competition studies using high-affinity 5-HT1B compounds performed in the presence of 100 nM 8-OH-DPAT (to block 5-HT1A receptors) indicated that approximately 26% of all specific, high-affinity [3H]5-HT binding to spinal cord synaptosomes was to 5-HT1B receptors. [3H]5-HT competition studies performed in the presence of 100 nM 8-OH-DPAT and 10 nM RU 24969 (to block 5-HT1A and 5-HT1B receptors, respectively) indicated that the remaining 49% of [3H]5-HT binding did not possess the pharmacologic profile previous reported for 5-HT1C, 5-HT1D, 5-HT1E, 5-HT2, or 5-HT3 receptors. This residual 49% of [3H]5-HT binding to spinal cord synaptosomes observed in the presence of 100 nM 8-OH-DPAT and 10 nM RU 24969 (subsequently referred to as "5-HT1S") displayed high affinity and saturability (KD = 4.7 nM) in association/dissociation and saturation experiments. Addition of 300 microM GTP or the nonhydrolyzable form of GTP, 5'-guanylylimidodiphosphate, inhibited [3H]5-HT binding to 5-HT1S receptors in saturation experiments by 35 and 57%, respectively, whereas ATP was without effect.(ABSTRACT TRUNCATED AT 250 WORDS)     

Water stress in Eucalyptus pauciflora: comparison of effects on stomatal conductance with effects on the mesophyll capacity for photosynthesis,and investigation of a possible involvement of photoinhibition

Seedlings of Eucalyptus pauciflora Sieb. ex Spreng., grown in 4-1 pots, were stressed by withholding water while relationships between net assimilation rate (A) and intercellular partial pressure of CO₂ (p_i) in selected leaves were obtained repeatedly throughout the stress cycle. Water stress at first caused stomatal closure without any decline in the A(p_i) relationship. As stress became more severe, the A(p_i) relationship was affected as well. This always affected assimilation rate at both high and low intercellular partial pressures of CO₂. It was then tested whether water-stressed leaves were more prone to photoinhibition than unstressed ones. Plants were water-stressed while at the same time subjected to strong photon flux area density (2000 mol quanta·m^-2·s^-1). A possible light-induced inhibition was assessed by comparing quantum yields of photosynthesis with light directed onto one or the other surface of the leaf. A decline in quantum yield was observed, and the decline on the previously irradiated side was more pronounced than on the previously shaded side, but the effect was small and disappeared entirely within 1 d of rewatering the plants. It is concluded that photoinhibition can play a role, but not an important one, in the effect of water stress on the A(p_i) relationship in leaves of E. pauciflora.Abbreviations and symbols RuBP ribulose-1,5-bisphosphate - A net assimilation rate - p_i intercellular partial pressure of CO₂ - quantum yield of photosynthesis (net assimilation or RuBP-regeneration rate) - w difference in water content between air saturated at leaf temperature and the actual vapor content of the air, expressed as mole fraction     

Differentiation antigens in human T-cell activation: Evidence that anti-VH antibodies react with a membrane structure on human T Lymphocytes distinct from the antigens detected by monoclonal antibodies of the OKT and Leu series

The effect of a panel of monoclonal antibodies and heteroantibodies on T-cell proliferation in various assay systems has been examined. The antibodies tested were directed against T-cell differentiation antigens, HLA-DR antigens, and structures defined by an anti-human V_H antiserum. As the test cell system highly purified subpopulations of T-cell growth factor (TCGF)-dependent T-cell lines activated either by mitogen or antigen were used. A survey of the data indicates the following: (1) Mitogenic and antigenic triggering of T lymphocytes are mediated through partly different membrane structures. (2) Antigenic stimulation by purified protein derivative (PPD) as well as polyclonal activation induced by OKT3/anti-Leu 4 monoclonal antibodies can be inhibited by heteroantibodies raised against human immunoglobulin V_H fragments thus pointing to a possible connection between the antigens detected by these antisera. (3) There does not seem to be differences between the two major subpopulations of T lymphocytes (i.e., helper/inducer and suppressor/cytotoxic cells) as to how they respond to antigens or mitogens in the investigated assay systems. (4) A clear distinction was found between T blasts specific for PPD and allogeneic cells as compared to cytotoxic T cells (CTL), as the T4 and T8 antigens seem to be functionally important for antigen recognition among CTL but not for the blasts proliferating in response to PPD and allogeneic cells. (5) An inhibitory effect of OKT3/anti-Leu 4, OKIal, and anti-HLA-DR on TCGF-dependent growth was detected, possibly indicating a steric relationship between these antigens and TCGF receptors on mitogen-induced T blasts. (6) Soluble factors obtained after incubating adherent cells with OKIal and anti-HLA-DR antibodies seemed to have an inhibitory effect on overall T-cell proliferation stressing the importance of studying the T-cell activation process at different levels in these kinds of experiments. (7) The results further suggest a complexity in the build up of antigen receptors on the various T-effector cells, perhaps also involving receptors for growth factors, HLA-DR antigens, and receptors for the latter.