Effects of high calcium levels on the disturbed extrusion of the second polar body during in vitro fertilization in C3H/He mouse substrains.

We previously reported that high concentrations (≥3.42 mM) of calcium during in vitro fertilization (IVF) disturbed the extrusion of the second polar body (PBII) in C3H/He inbred mice. In this study, the substrain specificity of this phenomenon was examined under 1.71-6.84 mM calcium concentration in ova from six C3H/He mouse commercially available substrains in Japan. PBII extrusion in ova from J substrains was not affected by calcium concentrations (<10% at any calcium level), but was grossly disturbed at high calcium levels in the ova of other substrains. This result has practical applications for the efficient production of normal zygotes by IVF, therefore contributing to the reduction in the numbers of donor animals for further zygote or embryo manipulation. Care must be taken in choosing IVF medium for particular strains and substrains.

Importance of absorbable surgical sutures for the prevention of stitch abscess after surgery in patients with oral squamous cell carcinoma.

BACKGROUND: To elucidate the significance of absorbable surgical sutures in the occurrence of stitch abscess after surgery in patients with oral squamous cell carcinoma (SCC). MATERIAL AND METHODS: The subjects were 251 patients who underwent excision and/or reconstruction and/or neck dissection for oral SCC using absorbable surgical sutures. Detection rates and characteristics of patients with stitch abscess were retrospectively evaluated by comparing between our present and previous data. RESULTS: There was only one stitch abscess among the 251 patients. A significant difference in the incidence of stitch abscess was found between the present data and our previous data. Of course, no significant correlations were found between the occurrence of stitch abscess using absorbable surgical sutures and the various factors seen in our previous analysis. CONCLUSIONS: A complete switch of surgical sutures from silk to absorbable surgical sutures is needed for surgery in patients with oral SCC.

Alterations in 18F-FDG accumulation into neck-related muscles after neck dissection for patients with oral cancers.

BACKGROUND: 18F-fluoro-2-deoxy-D-glucose (18F-FDG) accumulations are commonly seen in the neck-related muscles of the surgical and non-surgical sides after surgery with neck dissection (ND) for oral cancers, which leads to radiologists having difficulty in diagnosing the lesions. To examine the alterations in 18F-FDG accumulation in neck-related muscles of patients after ND for oral cancer. MATERIAL AND METHODS: 18F-FDG accumulations on positron emission tomography (PET)-computed tomography (CT) in neck-related muscles were retrospectively analyzed after surgical dissection of cervical lymph nodes in oral cancers. RESULTS: According to the extent of ND of cervical lymph nodes, the rate of patients with 18F-FDG-PET-positive areas increased in the trapezius, sternocleidomastoid, and posterior neck muscles of the surgical and/or non-surgical sides. In addition, SUVmax of 18F-FDG-PET-positive areas in the trapezius and sternocleidomastoid muscles were increased according to the extent of the ND. CONCLUSIONS: In evaluating 18F-FDG accumulations after ND for oral cancers, we should pay attention to the 18F-FDG distributions in neck-related muscles including the non-surgical side as false-positive findings.

Antibody-like proteins that capture and neutralize SARS-CoV-2.

To combat severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) and any unknown emerging pathogens in the future, the development of a rapid and effective method to generate high-affinity antibodies or antibody-like proteins is of critical importance. We here report high-speed in vitro selection of multiple high-affinity antibody-like proteins against various targets including the SARS-CoV-2 spike protein. The sequences of monobodies against the SARS-CoV-2 spike protein were successfully procured within only 4 days. Furthermore, the obtained monobody efficiently captured SARS-CoV-2 particles from the nasal swab samples of patients and exhibited a high neutralizing activity against SARS-CoV-2 infection (half-maximal inhibitory concentration, 0.5 nanomolar). High-speed in vitro selection of antibody-like proteins is a promising method for rapid development of a detection method for, and of a neutralizing protein against, a virus responsible for an ongoing, and possibly a future, pandemic.

Predicting the therapeutic effect of carbamazepine in trigeminal neuralgia by analysis of neurovascular compression utilizing magnetic resonance cisternography.

The aim of this study was to determine whether the evaluation of neurovascular compression (NVC) using new criteria on magnetic resonance (MR) cisternography improves the prediction of the curative effects of carbamazepine (CBZ) in trigeminal neuralgia (TN). In this study, a new analysis of NVC using MR cisternography was performed retrospectively for 280 patients with clinical signs and symptoms suggesting TN. The new analysis examined whether the site of the NVC was less than 3mm from the edge of the exit point and within the first third of the root entry zone on MR cisternography. Prediction of the curative effects of CBZ using the new evaluation of distance to the NVC improved on the predictive ability of the previous method of calculating the NVC volume (Mantel-Haenszel coefficient, P<0.01). In particular, initial treatment with CBZ 100mg/day for 2 weeks appeared more effective for patients with NVC volumes of ≤5mm3 plus a distance to the NVC of ≤3mm, than for those with NVC volumes >5mm3. The evaluation of NVC on MR cisternography using this new approach appears to be more useful than the previous method for predicting the initial treatment response in patients with TN.

A pedigree of amyotrophic chorea with acanthocytosis.

Several pedigrees of which some members showed a clinical syndrome consisting of mental changes, choreatic involuntary movements, limb muscles atrophy, and acanthocytosis have been reported in the United States and the United Kingdom. Such a case and some of the family members who had such abnormalities as acanthocytosis, hypo-beta-lipoproteinemia, convulsions, and confusion was observed. Results of biochemical analysis of catecholamines and their metabolites in CSF and urine showed an elevated value of norepinephrine in CSF and increased urinary secretion of DOPAC. The authors propose to designate this syndrome an amyotrophic chorea with acanthocytosis.

Behavioural and neurochemical effects of OPC-14597, a novel antipsychotic drug, on dopaminergic mechanisms in rat brain.

OPC-14597 is a new antipsychotic drug with a unique pharmacological profile. In a behavioural study in rats OPC-14597 did not show cataleptogenic activity even at the highest dose (40 mg/kg, i.p.), whereas it antagonized apomorphine-induced stereotypy dose-dependently (0.5-40 mg/kg). In vivo microdialysis showed that extracellular dopamine (DA) in the striatum was decreased significantly after OPC-14597 administration at higher doses of 10 and 40 mg/kg. Similar results were obtained in extracellular dopamine concentration in the frontal cortex, although the changes in DOPAC and HVA concentrations were smaller than those in the striatum. OPC-14597 also antagonized DA increase induced by the DA autoreceptor antagonist (+)-AJ76. These results that OPC-14597 acts either as an antagonist at postsynaptic dopamine receptors or as an agonist at presynaptic dopamine autoreceptors.

A substance P-containing pathway from the hypothalamic ventromedial nucleus to the medial preoptic area of the rat: an immunohistochemical analysis.

The destruction of the hypothalamic ventromedial nucleus, which contains a group of substance P-like immunoreactive neurons, resulted in a marked ipsilateral reduction of these fibers in the medial preoptic area. To test if and to what extent the substance P-like immunoreactive neurons in the ventromedial nucleus project to the medial preoptic area, we applied a sensitive double-labeling method capable of detecting substance P-like immunoreactivity in neurons retrogradely labeled with biotin-wheat germ agglutinin following injection of the tracer in the medial preoptic area. The appearance of many double-labeled cells in the hypothalamic ventromedial nucleus provides strong evidence for the existence of a prominent substance P containing pathway from the ventromedial nucleus to the medial preoptic area. A few doubled-labeled cells were also seen in the lateral hypothalamus, which therefore seems to be an additional source of substance P-like immunoreactive fibers in the medial preoptic area.

Ontogeny of the calcitonin gene-related peptide in the nervous system of rat brain stem: an immunohistochemical analysis.

The ontogeny of the calcitonin gene-related peptide in the neuron system of the rat brain stem was investigated by means of the indirect immunofluorescence technique. Calcitonin gene-related peptide-like immunoreactivity was first detected in the fibers of the nucleus of spinal tract trigeminal nerve on gestational day 18, and thereafter appeared gradually in various brain stem areas such as in the fibers of the solitary tract, gracile nucleus, cuneate nucleus, inferior colliculus, superior colliculus, medial geniculate nucleus and in the neurons of the hypoglossal nucleus, facial nucleus, superior olive, parabrachial area, superior colliculus and peripeduncular nucleus. In colchicine-untreated animals, the immunoreactive fibers increased in number and reached adult level by postnatal day 14, whereas the number of cells reached a maximum between postnatal days 2 and 6 and then decreased in number and immunoreactivity or disappeared, except in some areas such as the superior olive and peripeduncular nucleus, which showed the same immunoreactivity as for adult animals. With colchicine treatment, calcitonin gene-related peptide-like immunoreactive cells were found in more areas of the brain stem such as the abducens nucleus, parabigeminal nucleus, principal oculomotor nucleus, trochlear nucleus and central gray, along with the nuclei which had shown calcitonin gene-related peptide immunoreactivity in the untreated animals. However, the neurons in the inferior olive showed a different ontogenetical pattern of calcitonin gene-related peptide of immunoreactivity. Immunoreactivity disappeared completely by postnatal day 21 in both colchicine-untreated and -treated animals.

Influence of age on NMDA receptor complex in rat brain studied by in vitro autoradiography.

N-methyl-D-aspartate (NMDA) receptors are known to play an important role in learning and memory and to be involved in neuron cell death accompanying cerebral ischemia, seizures, and Alzheimer's disease. The NMDA receptor complex has been considered to consist of an L-glutamate recognition site, a strychnine-insensitive glycine modulatory site, and a voltage-dependent cation channel. In the present study, effects of age on an L-glutamate recognition site and a glycine site were examined in rat brain by quantitative in vitro autoradiography with [3H]-CPP and [3H]-glycine. Both [3H]-glycine and [3H]-CPP binding sites were most abundant in the hippocampus and cerebral cortex, and they showed a similar distribution pattern throughout the brain. [3H]-glycine binding sites were severely decreased in the telencephalic regions, including the hippocampus and cerebral cortex, in aged brain. Conversely, [3H]-CPP binding sites were well preserved in these brain areas. In the mid-brain regions and cerebellum, neither [3H]-glycine nor [3H]-CPP binding sites changed in the aged brain. Our results indicate that within the NMDA receptor complex, glycine receptors are primarily affected in the aging process.

The primordial to primary follicle transition.

The mechanisms that regulate the gradual exit of ovarian follicles from the non-growing, primordial pool are very poorly understood. A better understanding of the signals that initiate follicular growth in mammals, and of the conditions necessary for sustained growth of early preantral follicles in vitro, could have practical implications for contraception, alleviation of infertility, and regulation of the rate of follicle depletion (menopause). Our laboratory has developed two experimental systems that can be used to study factors involved in the activation of primordial follicles. In the first experimental system, small pieces of ovarian cortex, containing mostly primordial follicles, are isolated from fetal ovaries of cattle or baboons and cultured in serum-free medium. Under these conditions most primordial follicles become activated between 12 and 24 h of culture; their granulosa cells change shape, from flattened to cuboidal, and begin to express proliferating cell nuclear antigen (PCNA). During 7 days in culture, the newly-formed primary follicles and their oocytes increase significantly in diameter. This wholesale 'spontaneous' activation in serum-free medium is quite different from the much more gradual exit of primordial follicles from the resting pool that occurs in vivo and suggests that primordial follicles in vivo may be subject to a tonic inhibition of growth initiation or, alternatively, that some aspect(s) of the environment in vitro stimulates growth initiation. Recently we developed a second experimental system for studying activation of primordial follicles. Pieces of ovarian cortex from bovine or baboon fetuses were grafted beneath the developing chorioallantoic membrane (CAM) of 6-day-old chick embryos, a site known to support xenografted tissues. The cortical pieces were rapidly vascularized and histological analysis of pieces recovered after 2, 4, 7, or 10 days 'in ovo' revealed no increase in the number of primary follicles and maintenance of original numbers of primordial follicles. Therefore, grafting ovarian cortical pieces beneath the chick CAM provides an experimental system in which follicles remain at the primordial stage in a readily accessible environment and which, thus, may be used to study potential regulators of the initiation of follicle growth. The results suggest that vascularization of isolated pieces of ovarian cortex provides conditions that maintain follicular quiescence, whereas culture in vitro allows unrestrained activation of primordial follicles. Future studies with and comparisons of the in vitro and in ovo models may provide new insight into the mechanisms that regulate the primordial to primary follicle transition.

Treatment by pacemaker in familial amyloid polyneuropathy.

Amyloid deposits often involve the heart and cause disturbances in conduction and impulse formation in patients with familial amyloid polyneuropathy (FAP). Seven patients with FAP required pacemaker treatment during eight years. The most frequent bradyarrhythmias requiring pacing were sinus node dysfunction with junctional failure. Our seven patients had attacks and symptoms of bradyarrhythmias. A pacemaker relieved symptoms of bradycardias with recurrence of dislocation of electrode, exit block, and relatively high threshold. However, pacing did not improve the ultimate prognosis of FAP, because of progressive inanition of FAP. In our series, pacing should have been started earlier before advanced stage according to the ECG findings as in other diseases with bradycardias.

Modulation of dopamine D1 receptor binding by neurotensin in the rat striatum.

The effect of neurotensin on binding characteristics of dopamine D1 receptors was examined in the rat striatal membranes through radioreceptor assay. Neurotensin or its analogs were added to incubation medium of[(3)H]SCH 23390 saturation or dopamine/[(3)H]SCH 23390 inhibition experimental systems. Neurotensin did not modulate D1 antagonist binding but converted a part of D1 agonist high affinity binding sites to a low affinity state. Neurotensin(8-13) had the same potency as neurotensin itself, whereas neurotensin(1-8) had only weak activity in modulating D1 agonist binding. GTP and neurotensin had the same effect on D1 agonist binding. However, when both neurotensin and GTP were added, the result was the same as with either alone. These data suggest that neurotensin modulates the functional state of D1 receptors probably via a GTP binding protein in the rat striatum.

Differential expression of c-fos mRNA in rat prefrontal cortex, striatum, N. accumbens and lateral septum after typical and atypical antipsychotics: an in situ hybridization study.

The regional difference in the expression of c-fos mRNA induced by typical and atypical antipsychotics was determined in prefrontal cortex, striatum, N. accumbens and lateral septum in rats by in situ hybridization. Two typical antipsychotics, haloperidol (2 mg/kg) and fluphenazine (2 mg/kg), and three atypical antipsychotics, (-)sulpiride (100 mg/kg), clozapine (20 mg/kg) and OPC-14597 (40 mg/kg), were used. Brains were fixed with 4% paraformaldehyde 45 min after drug administration (i.p.). Brain sections of 30 microns-thickness were made in a cryostat and hybridized with 35S-labelled for c-fos oligonucleotide probe. These sections were apposed to X-ray films and the autoradiograms were semi-quantitatively analysed by computer-assisted densitometry. All antipsychotics used increased c-fos mRNA expression in N. accumbens shell, a region of the forebrain associated with limbic systems. On the other hand, two typical antipsychotics (haloperidol and fluphenazine) that cause a high incidence of acute motor side effects increased the expression of c-fos mRNA in the dorsolateral striatum, an extrapyramidal region primarily involved in motor control. Only clozapine induced c-fos mRNA in the medial prefrontal cortex and lateral septum. These results strongly suggest that the shell region of N. accumbens may be a common site of therapeutic action of antipsychotics.

Studies on biological actions of dimethyl sulfoxide in familial amyloidosis.

DMSO was therapeutically administered to patients with FAP and in about half of the patients there was some clinical improvement. Urinary proteins were analyzed biochemically and immunochemically before and after DMSO administration in seven cases of amyloidosis. As the results, increased excretion of various proteins of different molecular weights in the urine was observed depending on cases and examined organs. The in vitro effects of DMSO on amyloid proteins were examined. DMSO-degraded amyloid proteins showed void-volume materials and lower molecular weight components on Sephadex G column elution profiles as did guanidine-degraded amyloid protein. Among various denaturating or reducing agents, DMSO is the least potent in dissolving amyloid fibrils into prealbumin-related proteins.

NG-monomethyl-L-arginine, an inhibitor of nitric oxide synthase, increases extracellular GABA in the striatum of the freely moving rat.

We investigated the effect of an NO synthase inhibitor, NG-monomethyl-L-arginine (L-NMMA), on the levels of endogenous GABA in the rat striatum using in vivo microdialysis. Rats were perfused with the artificial CSF containing L-NMMA (0.1, 0.3 and 1.0 mM) or its inactive isomer D-NMMA (1.0 mM) for 1 h. Infusion of L-NMMA, but not its D-isomer, dose-dependently increased GABA concentration. Co-infusion with tetrodotoxin (1 microM) did not antagonize the increase of GABA induced by L-NMMA. These results show that decreased NO activity enhances GABA release even in the absence of depolarization of GABA neurones. We conclude that NO may be directly acting on GABA nerve terminals and tonically inhibiting GABA release or synthesis under basal conditions.

Coexistence of substance P and neurotensin-like peptides in single neurons of the rat hypothalamus.

The coexistence of substance P with neurotensin-like immunoreactivity in certain neurons of the hypothalamus were demonstrated by the double immunofluorescence method. Substance P and neurotensin-like immunoreactivity coexisted within single neurons of some hypothalamic areas such as the medial preoptic area, perifornical area, anterior hypothalamic area, lateral hypothalamic area, periventricular nucleus and posterior hypothalamic nucleus, although they did not coexist in the majority of immunoreactive cells.

The colocalization of substance P- and somatostatin-like peptides in neurons of the entopeduncular nucleus of rats.

The colocalization of substance P-like and somatostatin-like immunoreactivity in cell bodies was investigated in the entopeduncular nucleus of rats by a double immunofluorescence method using species specific antibodies. Most of the substance P-like immunoreactive cells were also positive to somatostatin and mainly seen in the rostral to middle region of the entopeduncular nucleus. Therefore it is suggested that double-labeled neurons in the entopeduncular nucleus project to the lateral habenular nucleus which is involved in the limbic system, since the rostral portion of the entopeduncular nucleus has been shown to project to the lateral habenular nucleus.

Atrial natriuretic polypeptide-like immunoreactivity in the rat pituitary: light and electron microscopic studies.

Immunoreactive atrial natriuretic polypeptide (ANP) was investigated in the pituitary of rats by light and electron microscopy using the indirect immunofluorescence and peroxidase-antiperoxidase techniques. ANP-like immunoreactivity was present in 30-35% of anterior pituitary cells. These cells have two types of secretory granules being characteristic of rat gonadotrophin-storing granules, and were usually adjacent to the capillary endothelium. The results of this study suggest the co-occurrence of ANP and gonadotrophins in the anterior pituitary cells.

Immunoreactive atrial natriuretic polypeptides in the adrenal medulla and sympathetic ganglia.

Atrial natriuretic polypeptide (ANP)-like immunoreactivity was found in the rat adrenal gland by using indirect immunofluorescence and peroxidase-antiperoxidase techniques. ANP-like immunostaining was present in most of chromaffin cells with varying degrees of immunoreactivity. The majority of medullary cells displayed very intense immunostaining, and several clusters revealed weaker immunostaining. No staining was found in the adrenal cortex or in the nerve fibers in this organ. In the consecutive sections treated for dopamine-beta-hydroxylase (DBH), apparently all medullary cells had intense immunofluorescence for DBH and its distribution pattern was very similar to that for ANP-like immunoreactivity. While phenylethanolamine N-methyltransferase (PNMT) immunoreactive cells largely corresponded to the intensely stained ANP-like immunoreactive cells, suggesting that adrenaline cells contained a large amount of ANP-like substance, noradrenaline cells contained a smaller amount of this substance than adrenaline cells. Ultrastructural study showed that end-products due to the immunoreaction with the ANP antiserum were primarily associating with chromaffin granules. In addition, the presence of ANP-like immunoreactivity was investigated in several sympathetic ganglia of the rat. No principal ganglion cells were ANP-positive, whereas a few small intensely fluorescent (SIF) cells were ANP-immunoreactive. The present findings suggest that catecholamines coexist with ANP which has a natriuretic and vasodilating effect, in adrenal medullary cells and SIF cells in several rat sympathetic ganglia, but not in principal ganglion cells.