Similarities between the relaxations induced by vasoactive intestinal peptide and by stimulation of the non-adrenergic non-cholinergic neurons in the rat stomach

Summary The characteristics of the non-adrenergic, noncholinergic inhibitory response of the rat stomach fundus to transmural nerve stimulation were compared with the relaxation induced by vasoactive intestinal polypeptide (VIP). Treatment with -chymotrypsin (5 U/ml) or VIP antiserum (1:200) significantly reduced the relaxation induced by transmural nerve stimulation at 30 Hz, indicating that the possible transmitter in the non-adrenergic, non-cholinergic nerves is a peptide and may be VIP or a closely related peptide. VIP was able to relax, fully and dose-dependently, the stomach fundus that had previously been constricted by treatment with 10^–6 M serotonin, and the IC₅₀ value for VIP was 2.4 × 10^–9 M. VIP elevated levels of cyclic AMP in a dose-dependent manner and the EC₅₀ value was 2.8 × 10^–9 M in the presence of 10^–6 M atropine and 10^–6 M guanethidine. The stomach fundus was relaxed by transmural nerve stimulation (30 Hz, 50 mA) and transmural nerve stimulation also caused production of cyclic AMP in the rat stomach in the presence of atropine and guanethidine. The basal level of cyclic AMP in the stomach was 8.7 ± 0.26 pmole/mg protein. When transmural nerve stimulation was applied for 5 min, the contraction of the stomach, induced by 10^–6 M serotonin, was inhibited by 54% in the presence of atropine and guanethidine and the level of cyclic AMP was increased to 13.0 ± 0.73 pmol/mg protein. Apamin inhibited the transmural nerve stimulation-induced relaxation and shifted the dose-response curve for VIP to the right. These results suggest that one of the putative neurotransmitter from non-adrenergic, non-cholinergic nerves in the rat stomach is VIP and that VIP-induced relaxation may be mediated by the production of cyclic AMP and by the opening of apamin-sensitive K⁺-channels.Send offprint requests to K. Kamata at the above address     

Catecholamine alterations in experimental hydrocephalus

Experimental hydrocephalus was induced in rats by intracisternal injection of kaolin suspension. The amounts of norepinephrine and dopamine were determined in the whole brain and specific brain regions at 1 week (acute phase) and 4 weeks (chronic phase). The turnover of catecholamine, an index of the activity of catecholamine-containing neurons, was determined by measuring the decrease in catechlamine contents 2 h after intraperitoneal injection of -methyl-p-tyrosine (250 mg/kg), an inhibitor of tyrosine hydroxylase. We observed that the catecholamine contents in kaolin-induced hydrocephalus were not significantly different from control values. Following injection of -methyl-p-tyrosine, there was decrease in levels of catecholamines in both control and hydrocephalic rats. This decrease was, however, significantly less in induced hydrocephalus than in control animals. This result suggested that in hydrocephalus, the activities of norepinephrinergic and dopaminergic neurons are reduced.     

“Skull punch” for craniotomy in neonates and infants

We have designed a new instrument that we have called a skull punch, which is useful and safe for making a burr hole for a purely osteoplastic craniotomy in neonates and infants. The instrument is described in detail.     

Evidence for a role of basal ganglia in the regulation of rapid eye movement sleep by electrical and chemical stimulation for the pedunculopontine tegmental nucleus and the substantia nigra pars reticulata in decerebrate cats

The present study was to determine how afferents from the substantia nigra pars reticulata (SNr) of the basal ganglia to the pedunculopontine tegmental nucleus (PPN) in the brainstem could contribute to the control of behavioral states. We used anesthetized and acutely decerebrated cats (n=22). Repetitive electrical stimulation (10-100 Hz, 20-50 microA, for 4-20 s) to the ventrolateral part of the PPN produced rapid eye movement (REM) associated with a suppression of postural muscle tone (REM with atonia). Although repetitive electrical stimuli (10-200 Hz, 10-60 microA, for 5-20 s) delivered to the dorsolateral part of the SNr did not evoke eye movements or muscular tonus in baseline conditions, it altered the PPN-induced REM with atonia. The following three types of effects were induced: (1) attenuation of the REM with atonia; (2) attenuation of muscular atonia without changes in REM (REM without atonia); and (3) attenuation of only REM. The optimal stimulus sites for these effects were intermingled within the lateral part of the SNr. The PPN-induced REM with atonia was abolished by an injection into the PPN of muscimol (1-15 mM, 0.1-0.25 microl), a GABAA receptor agonist, but not altered by an injection of baclofen (1-10 mM, 0.1-0.25 microl), a GABAB receptor agonist. Moreover, an injection of bicuculline (1-15 mM, 0.1-0.25 microl), a GABAA receptor antagonist, into the PPN, resulted in REM with atonia. On the other hand, an injection of muscimol into the dorsolateral part of the SNr (1-15 mM, 0.1-0.25 microl) induced REM with atonia, which was in turn eliminated by a further injection of muscimol into the PPN (5-10 mM, 0.2-0.25 microl). These results suggest that a GABAergic projection from the SNr to the PPN could be involved in the control of REM with atonia, signs which indicate REM sleep. An excessive GABAergic output from the basal ganglia to the PPN in parkinsonian patients may induce sleep disturbances, including a reduction of REM sleep periods and REM sleep behavioral disorders (REM without atonia).     

Regional difference in the appearance of apoptotic cell death in the ligamentum flavum of the human cervical spine

Ossification or calcification of the ligamentum flavum (LF) is relatively common in the middle and lower cervical, thoracic, and lumbar spine but extremely rare in the upper cervical region. This clinical fact suggests that there exist local factors promoting or preventing ossification or calcification of LF. However, little is known about the differences in the ultrastructure and cellular alterations of the LF between the different spinal levels, even in the cervical spine. With electron microscopy, we examined samples of LF collected surgically from the upper and lower cervical spine regions; we then studied the apoptotic appearance of ligament cells using a preferential labeling method. We found direct evidence of apoptosis of ligament cells in the LF. Apoptosis was more apparent in the upper region samples than in the lower region samples. The spaces around the normal fibroblasts were filled with thick collagen fibrils, but the collagen fibrils disappeared around the apoptotic bodies and thin fibrils were formed. The difference of the level of apoptosis may correlate to the ultrastructual difference of LF, and our data will benefit further investigations seeking to clarify the mechanism of various pathological conditions in the human LF.     

Establishment of an HIV cell-cell fusion assay by using two genetically modified HeLa cell lines and reporter gene

Infection of human cells with the human immunodeficiency virus type I (HIV-1) can be mimicked by a fusion process between cells expressing the HIV envelope protein (Env) and cells expressing both human CD4 together with the appropriate human chemokine receptors. In this study, a T-tropic HIV cell-cell fusion assay was established that utilized CD4, human CXCR4 and HIV NL4-3 gp160 as fusion components and a T7 polymerase-activated luciferase as a reporter system. The HeLa T4 cells used, expressed CD4 and CXCR4, and the applied HeLa KS386 cells expressed HIV NL4-3 gp160. By combining HeLa T4 cells with HeLa KS386 cells, an approximately about 100- to 300-fold increase in luciferase activity could be elicited relative to the control. The addition of anti-CD4 monoclonal antibody (Mab) (RPA-T4) or anti-CXCR4 Mab (12G5) in the assay significantly inhibited the fusion event; in contrast, an anti-CCR5 Mab (2D7) had no effect, indicating that the fusion assay was CD4 and CXCR4 dependent. In this report, fusion events could be monitored by both the luciferase reporter system and syncytia formation. Fusion events were monitored and compared using these two approaches. The luciferase reporter system was found to be more sensitive than syncytia formation. Moreover, compared with previous HIV fusion models, such as using recombinant vaccinia viruses, this system has several advantages, including simplicity and sensitivity. Finally, the system provides a powerful tool to study fusion mechanisms mediated by T-tropic HIV gp160, as well as to screen for fusion-blocking antibodies and antiviral agents.     

Electrical analogy of diastolic pressure difference between left atrium and ventricle

We proposed a mathematical model to describe the early filling process of the left ventricle and applied the model toin vivo experiments. The solution of a second-order differential equation indicated that the pressure difference between the left atrium and ventricle during ventricular filling (PD) could be explained by a transient response, i.e. decremental oscillation, in an LCR circuit. Thereafter, we analysed the sequence of PD during vagal stimulation with two catheter-tip manometers in 12 anaesthetised dogs and evaluated changes in the parameters of the system under various haemodynamic conditions. The values of ω_n and ξ were quite stable among beats within an episode of vagal stimulation, between episodes and even among dogs, despite the changes in haemodynamic variables. Pericardiotomy and partial discommunication of the mitral valve with the left ventricular free wall by cutting the mitral chordal tendons decreased ω_n and increased ζ, mainly because of the increase in C_LV. Occlusion of the coronary vascular beds with large numbers of microspheres increased ω_n and decreased ζ, mainly because of the decrease in C_LV. Mitral obstruction with an inflated balloon (increase in R) abolished the oscillatory changes and produced and exponential decay sequence of PD. In conclusion, both the logical and experimental approaches indicated that the sequence of PD could be considered as decremental oscillation in the LCR circuit and the parameters ω_n and ξ could be good indices of the diastolic property of the left ventricle     

Anatomical study of the obturator internus, gemelli and quadratus femoris muscles with special reference to their innervation

The manner of innervation of the obturator internus, superior and inferior gemelli, and the quadratus femoris in humans (101 pelvic halves) and in rhesus monkeys (Macaca mulatta: 8 pelvic halves) were investigated. In most specimens, the inferior gemellus originated from the lateral surface of the ischial tuberosity and also from the medial surface (intrapelvic origin) just beneath the obturator internus and was covered by the falciform process of the sacrotuberous ligament. The superior gemellus was frequently innervated by the nerve to the obturator internus and the nerve to the quadratus femoris (60.4%), and the inferior gemellus was innervated by the obturator internus nerve in two specimens. The quadratus femoris nerve originated from more cranial segments than the obturator internus nerve, however these nerves had various communication patterns inside and outside the muscles. According to the intramuscular nerve distribution, in some specimens the branches to the superior gemellus from the quadratus femoris nerve extended to the inferior gemellus, and the branches to the inferior gemellus were distributed to the obturator internus. The present findings revealed that the positional relationships among the branches to the obturator internus and gemelli muscles are relatively constant, although the branching patterns and innervation patterns were varied. The various patterns and routes are considered to reflect the variability of the differentiation patterns of the anlage of the muscles. A possible schematic model of the positional relationships between the muscles and the nerves is proposed.     

Effect of malnutritional rehabilitation on tuberculin reactivity and complement level in rats

The purpose of this study was to clarify the effect of differing nutritional states on various components of the immune system, especially on the interplay of the complement system and cell-mediated immunity. Malnutrition was induced in Sprague—Dawley rats by feeding them diets containing 5% protein or 0.5% protein as compared with 18% protein in the diet of the controls. Nutritional rehabilitation was achieved in some experimental groups by transferring those fed 0.5% protein diet to the 18% protein diet. Malnutrition was confirmed by weight changes, biochemical findings in the sera, haematological observations and histological observation of the liver, and rehabilitation was confirmed by body weight increase and changes in other measurements. In rats suffering malnutrition, the tuberculin skin reactivity was suppressed. After feeding the 0.5% protein diet for 8 weeks, all the rats showed negative tuberculin skin reactions. In the malnourished rats, including those fed with 0.5% protein, the serum complement level decreased but did not show any significant differences as compared with the well nourished control group. After 1 week of nutritional rehabilitation, the tuberculin reactivity of six out of ten rats remained negative and after 2 weeks, all rats showed positive tuberculin reactions. After 1 week of nutritional rehabilitation, all the rats showed a normal or higher serum complement level. At this stage, two of the tuberculin-negative rats showed significantly higher titre of serum complement than even the controls.