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Localization of glial cell antigens in the brains of young normal mice and the dysmyelinating mutant mice, jimpy and shiverer.

Tissue sections from the brains of normal, jimpy, and shiverer mice were immunostained by the peroxidase antiperoxidase method for carbonic anhydrase (CA) and the putative astrocytic "markers" glutamine synthetase (GS) and glial fibrillary acidic protein (GFAP). The cells in normal gray matter that immunostained with anti-CA and anti-GS were similar to one another in size and process elaboration. In the normal gray matter there were relatively few GFAP-positive astrocytes. When present, these cells resembled the CA- and GS-positive cells; however, the GFAP appeared to be concentrated in the astroglial processes, as distinguished from the cell bodies. Glial cell processes, immunostained for CA or GS, surrounded blood vessels and unstained neurons in the normal gray matter. The glial cells in shiverer gray matter were similar to those in the normal gray matter. When stained for GS or GFAP, the glial cells in the jimpy gray matter appeared to be somewhat hypertrophied, and when the glial cells in this mutant were stained for CA, the nuclei appeared to be swollen. It was concluded that some of the CA-positive cells in the gray matter of the normal and of each mutant mouse brain could be astrocytes. The patterns of immunostaining in the white matter emphasized the different complements of glial cells in the mutants. In the normal and shiverer mouse corpus callosum, CA, in particular, was detected only in the oligodendrocytes, their processes, and myelin. However, the data concerning the jimpy mouse suggested that the few CA-positive cells in the corpus callosum of that mutant could be astrocytes.

Animals↗

Transferrin in the central nervous system of the shiverer mouse myelin mutant.

Transferrin, the iron mobilization protein, and its mRNA are normally present in oligodendrocytes. Previous reports using myelin mutants have shown both a decrease in transferrin protein and mRNA when the oligodendrocyte population is compromised. In this study the shiverer mouse mutant in which the oligodendrocyte population is numerically normal, but has both quantitatively diminished and qualitatively abnormal myelin was used. This animal model was chosen to address the question whether expression of the transferrin message and/or protein correlated more closely to the number of oligodendrocytes (normal) or the amount of myelin (abnormally low). A 1/2 to 2/3 decrease in transferrin protein occurred in all brain regions examined except for the spinal cord in the shiverer group compared to both heterozygous littermates and wild type controls. Levels of transferrin transcripts in the brain are not affected by the shiverer mutation. These results taken with previous reports from this laboratory indicate that the presence of oligodendrocytes is a requirement for normal expression of transferrin mRNA in brain but is not sufficient for normal values of the protein. The level of Tf protein correlates more closely with the amount of myelin present than it does with the numbers of oligodendrocytes present. These data are consistent with previous reports from our laboratory that transferrin accumulation by oligodendrocytes is associated with myelin production by these cells. These data further suggest transferrin mRNA may be constitutively expressed by oligodendrocytes and that the protein expression is regulated at the level of translation.

Animals↗

Thermoregulation in the diabetic-obese (db/db) mouse. The role of non-shivering thermogenesis in energy balance.

1. Thermoregulation and non-shivering thermogenesis have been studied in the genetically diabetic obese (db/db) mouse. 2. At all environmental temperatures between 33 and 10 degrees C the body temperature of the diabetic mice was lower than that of the normal littermates, the difference varying from 1.1 degrees C at 33 degrees C to 4.5 degrees C at 10 degrees C. 3. At 4 degrees C the diabetic mice rapidly died (3.2h) of hypothermia while the normal mice maintained their body temperature within the normal range. 4. At 23 degrees C the diabetic animals exhibited a diurnal rhythm in body temperature which was similar in both phase and amplitude to the controls, but at every point throughout the 24h cycle the temperature of the mutants was lower by 1--2 degrees C. 5. The resting metabolic rate at thermoneutrality (33 degrees C) was higher per whole animal for the diabetics than for the normals. However, at temperatures below thermoneutrality the converse was observed; between 30 and 4 degrees C the RMR of the mutants was lower than the controls by approximately 25%. 6. The capacity for non-shivering thermogenesis in diabetic mice was only one-half that found in normal animals. 7. The diabetic mouse has abnormalities in thermoregulation and non-shivering thermogenesis which are similar to those found in the genetically obese (ob/ob) mouse. It is concluded that the high metabolic efficiency of the diabetic mouse, like that of the ob/ob mouse, can be explained by a reduced energy expenditure on thermoregulatory thermogenesis; this may represent a primary mechanism for the operation of the "thirfty genotype" associated with obesity and diabetes.

Animals↗

Prevention of postanesthetic shivering with intravenous administration of aspirin.

There have been many reports of postanesthetic shivering (PAS); however, the causes have not been defined clearly, and the reported methods of inhibiting PAS are not always available clinically. In the present study, we assessed the effect of the intravenous administration of aspirin on the prevention of PAS in 62 patients undergoing oral or maxillofacial surgery, anesthetized with enflurane-nitrous oxide. Thirty of the patients were randomly selected, and received intravenous aspirin DL-lysine 900 mg (equivalent to aspirin 495 mg) before the end of surgery. No significant differences were noted in the rectal temperatures between the group given aspirin and the control group. Shivering was observed in 17 of the 32 patients of control group. In contrast, shivering was observed in 5 of the 30 patients who received aspirin. This was a statistically significant difference ( P < 0.01). These data indicate that intravenous administration of aspirin significantly inhibited PAS. The finding suggests that PAS is related to prostaglandin synthesis or to the formation of derivatives of arachidonic acid, since aspirin inhibits both the synthesis of prostaglandins and the formation of derivatives of arachidonic acid.

Clinical Trial↗

5'-nucleotidase localization in the brains and spinal cords of adult normal and dysmyelinating mutant (shiverer) mice.

Immunocytochemical staining with the antibody against mouse liver 5'-nucleotidase revealed 5'-nucleotidase antigenicity in myelinated fibers in the brains and in myelinated fibers and some interfascicular oligodendroglia in the spinal cords of normal adult mice. Although the 5'-nucleotidase specific activity in adult shiverer mouse CNS tissue homogenates had been shown to be normal, immunocytochemical staining with anti-mouse-5'-nucleotidase could be demonstrated in CNS tissue sections from only 2 out of 10 of the mutant animals. In tissue from these animals the staining, which was relatively faint, was localized specifically to cell-bodies, usually arranged in rows, and to material oriented parallel to nerve fibers. This pattern of immunostaining with anti-5'-nucleotidase resembled the immunostaining with anti-carbonic anhydrase but not with anti-glial-fibrillary-acidic-protein. This suggested that the rows of cells were oligodendrocytes, not astrocytes, and that the material parallel to nerve fibers might consist of oligodendrocyte processes wrapped loosely around axons. The antibody against rat 5'-nucleotidase, as distinguished from mouse, immunostained only the blood vessels in the shiverer mouse CNS, a finding similar to a previous observation in the normal mouse CNS. From these findings it was inferred that the primary loci of 5'-nucleotidase in the shiverer mouse CNS were interfascicular oligodendrocytes, their processes, and blood vessels, and in the normal mouse CNS, the myelin in some tracts, the blood vessels, and some interfascicular oligodendrocytes.

5'-Nucleotidase↗

Elevated specific activity of 5'-nucleotidase in a spinal cord myelin fraction from shiverer mice. Comparison with other myelin-associated enzymes and myelin proteins.

Myelin partially purified from spinal cords of dysmyelinating mutant (shiverer) mice had almost three-fold the specific activity of 5'-nucleotidase found in the respective myelin fraction from normal mice. The specific activities of two other normally myelin-associated enzymes, 2',3'-cyclic nucleotide-3'-phosphohydrolase and carbonic anhydrase, were only slightly higher in the myelin membranes from shiverers, compared to those from controls. In the mutants, the three enzymes probably occur in oligodendrocyte processes. Hypothetically, the 5'-nucleotidase in the myelin sheath in shiverer and normal mice may be localized in specialized structures.

5'-Nucleotidase↗

Statistical diffusion tensor histology reveals regional dysmyelination effects in the shiverer mouse mutant.

Shiverer is an important model of central nervous system dysmyelination characterized by a deletion in the gene encoding myelin basic protein with relevance to human dysmyelinating and demyelinating diseases. Perfusion fixed brains from shiverer mutant (C3Fe.SWV Mbp(shi)/Mbp(shi)n = 6) and background control (C3HeB.FeJ, n = 6) mice were compared using contrast enhanced volumetric diffusion tensor magnetic resonance microscopy with a nominal isotropic spatial resolution of 80 mum. Images were accurately coregistered using non-linear warping allowing voxel-wise statistical parametric mapping of tensor invariant differences between control and shiverer groups. Highly significant differences in the tensor trace and both the axial and radial diffusivity were observed within the major white matter tracts and in the thalamus, midbrain, brainstem and cerebellar white matter, consistent with a high density of myelinated axons within these regions. The fractional anisotropy was found to be much less sensitive than the trace and eigenvalues to dysmyelination and associated microanatomic changes.

Animals↗

UCP1: the only protein able to mediate adaptive non-shivering thermogenesis and metabolic inefficiency.

The uniqueness of UCP1 (as compared to UCP2/UCP3) is evident from expression analysis and ablation studies. UCP1 expression is positively correlated with metabolic inefficiency, being increased by cold acclimation (in adults or perinatally) and overfeeding, and reduced in fasting and genetic obesity. Such a simple relationship is not observable for UCP2/UCP3. Studies with UCP1-ablated animals substantiate the unique role of UCP1: the phenomenon of adaptive adrenergic non-shivering thermogenesis in the intact animal is fully dependent on the presence of UCP1, and so is any kind of cold acclimation-recruited non-shivering thermogenesis; thus UCP2/UCP3 (or any other proteins or metabolic processes) cannot substitute for UCP1 physiologically, irrespective of their demonstrated ability to show uncoupling in reconstituted systems or when ectopically expressed. Norepinephrine-induced thermogenesis in brown-fat cells is absolutely dependent on UCP1, as is the uncoupled state and the recoupling by purine nucleotides in isolated brown-fat mitochondria. Although very high UCP2/UCP3 mRNA levels are observed in brown adipose tissue of UCP1-ablated mice, there is no indication that the isolated brown-fat mitochondria are uncoupled; thus, high expression of UCP2/UCP3 does not necessarily confer to the mitochondria of a tissue a propensity for being innately uncoupled. Whereas the thermogenic effect of fatty acids in brown-fat cells is fully UCP1-dependent, this is not the case in brown-fat mitochondria; this adds complexity to the issues concerning the mechanisms of UCP1 function and the pathway from beta(3)-adrenoceptor stimulation to UCP1 activation and thermogenesis. In addition to amino acid sequences conserved in all UCPs as part of the tripartite structure, all UCPs contain certain residues associated with nucleotide binding. However, conserved amongst all UCP1s so far sequenced, and without parallel in all UCP2/UCP3, are two sequences: 144SHLHGIKP and the C-terminal sequence RQTVDC(A/T)T; these sequences may therefore be essential for the unique thermogenic function of UCP1. The level of UCP1 in the organism is basically regulated at the transcriptional level (physiologically probably mainly through the beta(3)-adrenoceptor/CREB pathway), with influences from UCP1 mRNA stability and from the delay caused by translation. It is concluded that UCP1 is unique amongst the uncoupling proteins and is the only protein able to mediate adaptive non-shivering thermogenesis and the ensuing metabolic inefficiency.

Acclimatization↗

Chromosomal mapping of mouse myelin basic protein gene and structure and transcription of the partially deleted gene in shiverer mutant mice.

The gene for mouse myelin basic protein (MBP) was mapped to chromosome 18 by hybridization of cloned MBP probes to DNA from hamster-mouse hybrid cell lines, showing it to be linked to the shiverer mutation which causes abnormal CNS myelination. Genomic blotting experiments show that in the mutants five of six exons which constitute the wild-type gene have been deleted. In shiverer brains the steady state level of transcripts that initiate correctly at the 5' end of the remaining exon 1 is reduced 16-fold. These RNAs are not spliced correctly and are not efficiently polyadenylated. It is proposed that the partial deletion of the MBP gene is an important part of the shiverer lesion.

Animals↗

Transplantation of CNS fragments into the brain of shiverer mutant mice: extensive myelination by implanted oligodendrocytes. I. Immunohistochemical studies.

Solid fragments of olfactory bulb from new-born normal (B6CBA and C57BL6) mice were implanted into new-born shiverer (shi/shi) brains. The shiverer mouse being characterized by the absence of myelin basic protein (MBP), myelination due to implanted oligodendrocytes can be detected in the shiverer brain using an antiserum anti-MBP. Observation of sagittal sections of the host brains revealed very extensive areas of normal myelination from the level of the graft (rostral thalamus) up to the caudal brain (diencephalon, cerebellum, pons). Thus, oligodendrocytes contained in the implant migrate out of the graft over long distances in the host brain, before they differentiate and synthesize myelin. These results raise the question of the behaviour of oligodendrocytes in normal development.

Animals↗

Auditory brain stem response in neuropathological mutant mice (shiverer and reeler).

Auditory brain stem response (ABR) was investigated in shiverer mice and reeler mice. In shiverer mice, the wave latency after ABR wave III was prolonged and wave V disappeared. Although the interpeak latencies were also prolonged, there were some differences according to the site of the disorder. The threshold was found to be normal. In reeler mice, the findings on the wave latencies were inconsistent. Some of the mice showed no differences from the controls, some showed prolonged latencies, and some showed only wave I. The ABR findings on the shiverer mice were considered to be due to abnormal or poor myelin formation in the central nervous system. Abnormal ABR findings in the reeler mice are probably caused by a secondary change due to the abnormal cell positions resulting from mutation.

Animals↗

Elevated expression of type II Na+ channels in hypomyelinated axons of shiverer mouse brain.

Type I and type III Na+ channels are localized mainly in neuronal cell bodies in mouse brain. Type II channels are preferentially localized in unmyelinated fiber tracts but are not detectable in normally myelinated fibers. In shiverer mice, which lack compact myelin due to a defect in the myelin basic protein gene, elevated expression of type II Na+ channels was observed in the hypomyelinated axons of large-caliber fiber tracts such as the corpus callosum, internal capsule, fimbria, fornix, corpus medullare of the cerebellum, and nigrostriatal pathway by immunocytochemical analysis with subtype-specific antibodies. No difference was observed in the localization of type I and type III Na+ channels between wild-type and shiverer mice. These findings support the hypothesis that type II Na+ channels are preferentially localized in axons of brain neurons and suggest that their density and localization are regulated by myelination. The selective increase in the number of type II channels in hypomyelinated fiber tracts may contribute to the hyperexcitable phenotype of the shiverer mouse.

Animals↗

Clinical and pathologic findings in two draft horses with progressive muscle atrophy, neuromuscular weakness, and abnormal gait characteristic of shivers syndrome.

Two Belgian geldings, 4 and 14 years old, respectively, with muscle atrophy, weakness, and abnormal gait characteristic of severe advanced shivers were examined clinically and on necropsy. Neurologic examination revealed no evidence of ataxia, and the clinical diagnosis was neuromuscular weakness and shivers. Necropsies of both horses, including examination of pituitary, brain, spinal cord, spinal roots and ganglia, and peripheral nerves, revealed no gross or histologic abnormalities. Examination of multiple skeletal muscle specimens revealed chronic myopathic changes and periodic acid-Schiff positive, amylase-resistant inclusions within muscle fibers, characteristic of equine polysaccharide storage myopathy. It is suggested that underlying metabolic myopathy may be the cause of muscle weakness and cramping in horses with shivers.

Animal Feed↗

Factors which affect shivering in man during cold water immersion.

Six subjects were immersed in cold water (15.15 +/- 0.42 degrees C) and were asked to perform two tasks. Shivering elicited by the cold water immersion was attenuated and/or abolished by the mental arithmetic task and in some instances by a voluntary isometric contraction of forearm muscles. Some reasons for these results are discussed. Key words: Cold water immersion, mental arithmetic forearm isometric contraction, attenuation of shivering.

Cold Temperature↗

Thermoregulation and non-shivering thermogenesis in the genetically obese (ob/ob) mouse.

1. The capacity ofr thermoregulation and thermogenesis in lean and genetically obese (ob/ob) mice has been investigated. 2. At 4 degrees C ob/ob mice rapidly die of hypothermia, because of a reduced capacity for cold-induced thermogenesis, but the animals are able to survive if previously adapted to 12 degrees C. 3. At all environmental temperatures between 30 degrees C and 10 degrees C the body temperature of ob/ob mice is 2.0-2.5 degrees C below that of lean animals. This may be due to a lower "setting" for body temperature. 4. At 34 degrees C the oxygen consumption of obese mice is greater than that of the lean animals while at 30 degrees C it is similar. When the environmental temperature is below 30 degrees C the oxygen consumption of the lean mice is greater. The obese animals therefore expend less energy on thermoregulatory thermogenesis. 5. The capacity for non-shivering thermogenesis was measured in lean and obese mice by investigating the effect of an injection of L-nor-adrenaline (1000 microgram/kg body weight) on the metabolic rate at 31 degrees C. Non-shivering thermogenesis was reduced by one-half in the obese animals. 6. One cause of the obesity of the ob/ob mouse is its high metabolic efficiency. We suggest that this high metabolic efficiency is due, at least in part, to less energy being expended on thermoregulatory thermogenesis.

Animals↗

Prediction of shivering heat production from core and mean skin temperatures.

Prediction formulae of shivering metabolism (Mshiv) are critical to the development of models of thermoregulation for cold exposure, especially when the extrapolation of survival times is required. Many such formulae, however, have been calibrated with data that are limited in their range of core temperatures (Tc), seldom involving values of less than 36 degrees C. Certain recent studies of cold-water immersion have reported Tc as low as 33.25 degrees C. These data comprise measurements of Tc (esophageal) and mean skin temperature (Ts), and metabolism from 14 males [mean (SD); age = 28 (5) years; height = 1.78 (0.06) m; body mass = 77.7 (6.9) kg; body fat (BF) = 18.4 (4.5)%] during immersion in water as cold as 8 degrees C for up to 1 h and subsequent self-rewarming via shivering under dry blanketed conditions. The data contain 3343 observations with mean (SD) Tc and Ts of 35.92 (0.93) degrees C and 23.4 (8.9) degrees C, respectively, and have been used to re-examine the prediction of Mshiv. Rates of changes of these temperatures were not used in the analysis. The best fit of the formulae, which are essentially algebraic constructs with and without setpoints, are those with a quadratic expression involving Ts. This is consistent with the findings of Benzinger (1969) who demonstrated that the thermosensitivity of skin is parabolic downwards with temperature peaking near a value of 20 degrees C. Formulae that included a multiplicative interaction term between Tc and Ts did not predict as well. The best prediction using 37 degrees C and 33 degrees C as the Tc and Ts setpoints, respectively, was found with BF as an attenuation factor: Mshiv (W x m(-2)) = [155.5 x (37- Tc) + 47.0 x (33 - Ts) - 1.57 x (33 - Ts)2]/(%BF)(0.5).

Adult↗

Non-shivering thermogenesis during prostaglandin E1 fever in rats: role of the cerebral cortex.

We have tested the hypothesis that there is a role for the cerebral cortex in the control of non-shivering thermogenesis during fever induced by prostaglandin E1 (PGE1). While under urethan anesthesia, the firing rate of nerves innervating interscapular brown adipose tissue (IBAT), IBAT and colonic temperatures (TIBAT and Tc) and oxygen (O2) consumption were monitored during the fever from PGE1 injection (400 and 800 ng) in a lateral cerebral ventricle in controls and in functionally decorticated Sprague-Dawley rats. Rats were functionally decorticated by applying 3.3 M KCl solution on the frontal cortex which causes cortical spreading depression (CSD). Pyrogen injections caused dose-related increases in firing rate, TIBAT, Tc and O2 consumption and CSD reduced these enhancements. Our findings indicate that the cerebral cortex could be involved in the control of non-shivering thermogenesis during PGE1-induced febrile response.

Adipose Tissue, Brown↗

[Efficacy of ketanserin on postanesthetic shivering].

OBJECTIVE: To evaluate the clinical and electromyographic (EMG) effects of ketanserin (K), a serotoninergic receptor antagonist (5-HT2), on postoperative shivering (POS). STUDY DESIGN: Prospective, randomised, double-blind study. PATIENTS: Fifty ASA class 1 and 2 patients with major clinical postoperative tremor were studied. METHODS: POS was assessed clinically (0 = nil, 1 = moderate, 2 = severe). Inclusion criterion was a POS of 2 at admission in the recovery room. The mean arterial blood pressure, rectal temperature, SpO2 were recorded at admission (T0) and subsequently at T5, T10, T15, T30 and T60 minutes. Either 10 mg of K (n = 25) or a corresponding volume of a placebo (P) (n = 25) were intravenously injected. The EMG activity of the deltoid and quadriceps muscles was recorded continuously. Blood lactic acid concentration was measured at the end of POS. Results are expressed as mean +/- SEM. Parametric values were analysed with unpaired Student's t-test, and nonparametric values with chi 2 analysis. P < 0.05 was accepted. RESULTS: Demographic data, duration of anaesthesia, postoperative temperature, oxygen saturation, blood pressure and blood lactate concentration were similar between groups. The POS duration in the K group was significantly shorter than in the P group: 8.8 +/- 1.5 min and 15.5 +/- 1.5 min respectively (P < 0.01). The number of patients in the K group experiencing POS at T5 and T10 was significantly lower, when compared with those who had received the P (P < 0.05). CONCLUSION: At a dose of 10 mg, K administered in patients with POS during recovery, reduced significantly the duration and intensity of the shivering without noticeable side effects. This study suggests that this 5-HT2 antagonist is an efficient therapeutic tool for POS in adults.

Adult↗