Neurobehavioral deficits in mice lacking the erythrocyte membrane cytoskeletal protein 4.1

The erythrocyte membrane cytoskeletal protein 4.1 (4.1R) is a structural protein that confers stability and flexibility to erythrocytes via interactions with the cytoskeletal proteins spectrin and F-actin and with the band 3 and glycophorin C membrane proteins. Mutations in 4.1R can cause hereditary elliptocytosis, a disease characterized by a loss of the normal discoid morphology of erythrocytes, resulting in hemolytic anemia [1]. Different isoforms of the 4.1 protein have been identified in a wide variety of nonerythroid tissues by immunological methods [2-5]. The variation in molecular weight of these different 4.1 isoforms, which range from 30 to 210 kDa [6], has been attributed to complex alternative splicing of the 4.1R gene [7]. We recently identified two new 4.1 genes: one is generally expressed throughout the body (4. 1G) [8] and the other is expressed in central and peripheral neurons (4.1N) [9]. Here, we examined 4.1R expression by in situ hybridization analysis and found that 4.1R was selectively expressed in hematopoietic tissues and in specific neuronal populations. In the brain, high levels of 4.1R were discretely localized to granule cells in the cerebellum and dentate gyrus. We generated mice that lacked 4.1R expression; these mice had deficits in movement, coordination, balance and learning, in addition to the predicted hematological abnormalities. The neurobehavioral findings are consistent with the distribution of 4.1R in the brain, suggesting that 4.1R performs specific functions in the central nervous system.     

Anion exchanger 1 (band 3) is required to prevent erythrocyte membrane surface loss but not to form the membrane skeleton

The red blood cell (RBC) membrane protein AE1 provides high affinity binding sites for the membrane skeleton, a structure critical to RBC integrity. AE1 biosynthesis is postulated to be required for terminal erythropoiesis and membrane skeleton assembly. We used targeted mutagenesis to assess AE1 function in vivo. RBCs lacking AE1 spontaneously shed membrane vesicles and tubules, leading to severe spherocytosis and hemolysis, but the levels of the major skeleton components, the synthesis of spectrin in mutant erythroblasts, and skeletal architecture are normal or nearly normal. The results indicate that AE1 does not regulate RBC membrane skeleton assembly in vivo but is essential for membrane stability. We postulate that stabilization is achieved through AE1-lipid interactions and that loss of these interactions is a key pathogenic event in hereditary spherocytosis.     

Natural scenes are indeed preferred, but image quality might have the last word.

Numerous studies have shown that people prefer natural scenes over human-made scenes. Evolutionarily and neurologically based explanations for this preference have been proposed. This study examined the impact that image quality has on the preference for natural scenes. The authors also explored the influence of image quality on familiarity ratings and on how scenes are classified in a rapid, two-alternative, forced-choice task. Finally, the authors propose a framework for conceptualizing the nature of different image manipulation procedures, and how they influence aesthetic experiences. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Yes, I have heard about family psychotherapy—but opinions are no substitute for data.

E. Rudolph (see record 1982-26266-001) criticized an article by the present author (see record 1981-33210-001), saying that the existence and impact of family therapy was omitted from a review of the claims and progress of psychotherapy. It is argued that Rudolph's claim that family therapy represented a significant breakthrough in psychotherapy is unsupported by research data and that overall results of outcome in family therapy appear to be comparable to those for individual psychotherapy. Results of a study by S. L. Garfield and R. Kurtz (see record 1976-24639-001) show relatively little involvement of clinical psychologists with family therapy in terms of time devoted to professional activities. (4 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

SV40 T antigen directed by a powerful erythroid enhancer-promoter produced sarcomas and pancreatic tumors but not erythroid-specific tumors in transgenic mice

We have expressed the simian virus 40 (SV40) large T antigen oncogene in erythroid tissues of mice to test its ability to immortilize erythroid cells. A transgene construct was built in which the SV40 large T antigen structural gene was linked to erythroid-specific enhancer and promoter sequences. The enhancer employed was the human beta-globin family microlocus control region, and the promoter sequences were derived from the human beta-globin promoter. Transgenic mice were generated and they expressed T antigen in the bone marrow and spleen cells. Yet, no hematopoietic neoplasia arose in these mice. Instead, after a lag period of 2-6 months, the mice developed soft tissue sarcomas and pancreatic islet-cell tumors that expressed high levels of T antigen.     

Lipid metabolism and membrane composition are altered in the brains of type II diabetic mice

CBL/57 strain db/db mice exhibit type II (noninsulin-dependent) diabetes. The affected mice are markedly hyperinsulinemic, hyperglycemic, and hypercholesterolemic, and their serum K+ levels are decreased. The brains of the diabetic mice are significantly smaller than those of their lean, control littermates, but the protein concentration is normal. The low brain weight is accompanied by a loss of major fatty acid components within the whole brain, nerve endings, and mitochondrial membranes. Cholesterol levels are low in whole brain but are not significantly different from normal in the synaptosomal membranes. The phospholipid concentration is significantly decreased in whole brain homogenates, crude synaptosomal membranes, and crude mitochondrial membranes of the diabetic mice. In addition, the specific activities of membrane-bound synaptosomal acetylcholinesterase, Na+,K(+)-ATPase, and Mg(2+)-ATPase are decreased in crude synaptosomal membranes of the diabetic mice. The specific activities of carnitine palmitoyltransferase I and carnitine acetyltransferase are significantly increased in the crude mitochondrial fraction isolated from the brains of the type II diabetic mice, whereas the specific activity of pyruvate dehydrogenase complex is decreased. The specific activities of two other mitochondrial enzymes--monoamine oxidase B and citrate synthase--and a cytosolic enzyme--lactate dehydrogenase--are unaltered. The ability to synthesize cyclic AMP is markedly decreased in the brains of the diabetic mice. The concentrations of carnitine and of the amino acids, glutamate, aspartate, glutamine, and serine are unaltered, whereas glycine levels are significantly elevated in the brains of the db/db mice. The data suggest that in vivo the brains of the diabetic mice exhibit a decreased capacity for glucose oxidation and increased capacity for fatty acid oxidation. This hypothesis is supported by the finding that cerebral mitochondria isolated from the db/db mice oxidize [1-14C]palmitate to 14CO2 at a rate almost twice that of control mitochondria. The present findings emphasize the potentially serious alteration of brain metabolism in uncontrolled type II diabetes.     

Pseudomonas aeruginosa isolates from patients with cystic fibrosis have different beta-lactamase expression phenotypes but are homogeneous in the ampC-ampR genetic region

Pseudomonas aeruginosa isolates from 1 of 17 cystic fibrosis patients produced secondary beta-lactamase in addition to the ampC beta-lactamase. Isolates were grouped into three beta-lactamase expression phenotypes: (i) beta-lactam sensitive, low basal levels and inducible beta-lactamase production; (ii) beta-lactam resistant, moderate basal levels and hyperinducible beta-lactamase production; (iii) beta-lactam resistant, high basal levels and constitutive beta-lactamase production. Apart from a base substitution in the ampR-ampC intergenic region of an isolate with moderate-basal-level and hyperinducible beta-lactamase production, sensitive and resistant strains were identical in their ampC-ampR genetic regions. Thus, enhanced beta-lactamase expression is due to mutations in regulatory proteins other than AmpR.     

Most classes of dorsal root ganglion neurons are severely depleted but not absent in mice lacking neurotrophin-3

During development, many neurons in the dorsal root ganglia require neurotrophin-3 for survival. However, it is not known precisely which subpopulations of sensory neurons, other than the proprioceptive afferents, are neurotrophin-3 dependent in vivo. In this study, using a battery of neurochemical markers that label different subpopulations of dorsal root ganglion neurons, we found a widespread, about 60-65% loss of cells in most subpopulations in neurotrophin-3 deficient mice. Intermediate losses were found in the heterozygous mutant mice consistent with a gene dosage effect. In agreement with this, the cell size distribution between the homozygous mutant and wild type mice was virtually identical. The loss of small neurons containing calcitonin gene-related peptide, substance P and thiamine monophosphatase activity suggests that many unmyelinated primary afferents are also lost in the mutant animals. The fact that many different sensory neuron subpopulations are lost to the same extent in neurotrophin-3 deficient mice is consistent with the proposed early role of neurotrophin-3 during neurogenesis. Interestingly, calretinin immunoreactive neurons, which contribute a minor subpopulation, were not affected suggesting that neurotrophin-3 independent regulation of neurogenesis occurs in addition to prominent neurotrophin-3 dependent mechanisms.     

GluR-A-deficient mice display normal acquisition of a hippocampus-dependent spatial reference memory task but are impaired during spatial reversal.

Acquisition and reversal of a spatial discrimination were assessed in an appetitive, elevated plus-maze task in 4 groups of mice: knockout mice lacking the AMPA receptor subunit GluR-A (GluR1), wild-type controls, mice with cytotoxic hippocampal lesions, and controls that had undergone sham surgery. In agreement with previous studies using tasks such as the water maze, GluR-A-/- mice were unimpaired during acquisition of the spatial discrimination task, whereas performance in the hippocampal group remained at chance levels. In contrast to their performance during acquisition, the GluR-A-/- mice displayed a mild deficit during reversal of the spatial discrimination and were profoundly impaired during discrete trial, rewarded-alternation testing on the elevated T maze. The latter result suggests a short-term. flexible spatial working memory impairment in GluR-A-/- mice, which might also underlie their mild deficit during spatial reversal. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Cyclobutane thymine dimers with a disrupted phosphodiester bond are refractory to T4 endonuclease V digestion but have increased sensitivity to UvrABC nuclease

UV irradiation induces the dimerization of synthetic single-stranded, 80-mer oligonucleotides with self-complementary, alternating purine-pyrimidine sequences, and terminal 5'- and 3'-thymines; this process can be reversed by photoreactivation. The UV-induced 160-mers are sensitive to digestion by the restriction enzyme SnaBI, but monomers are insensitive to digestion, indicating that UV irradiation stabilizes the formation of double-stranded DNA. These results suggest that UV irradiation of these 80-mer oligonucleotide substrates induces the formation of a novel cyclobutane thymine dimer which lacks an intradimer phosphodiester bond (CPD*). This CPD*, linking the terminal thymines of two separate 80-mer molecules, is formed in a double-stranded DNA region created by self-annealing and intermolecular hybridization of the two 80-mer strands. We have found that these UV-induced CPD* in 160-mers are sensitive to cleavage by the nucleotide excision enzyme complex UvrABC nuclease, but resistant to cleavage by the cyclobutane pyrimidine dimer-specific enzyme T4 endonuclease V. However, pretreatment of the 160-mers with ligase reverses their sensitivity to these two enzymes, significantly reducing their susceptibility to cleavage by UvrABC nuclease but dramatically increasing their susceptibility to cleavage by T4 endonuclease. The biological significance of these findings is discussed.     

AMPA-receptors are involved in the expression of amphetamine-induced behavioural sensitisation, but not in the expression of amphetamine-induced conditioned activity in mice

We investigated the role of the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) receptor antagonist 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo[f]quinoxaline (NBQX) in the expression of amphetamine-induced behavioural sensitisation and amphetamine-induced conditioned activity in mice. Repeated weekly administration of amphetamine (0.375 mg/kg) for 7 weeks led to an increased locomotor response when challenged with amphetamine 1 week later. NBQX attenuated this increased response at doses (3 and 30 mg/kg) which had no effect on the acute locomotor response to amphetamine. In a separate experiment, mice given amphetamine (1 mg/kg) in a distinctive environment, showed an increased locomotor response within this environment following a subsequent saline administration. NBQX (5-20 mg/kg) had no effect on the expression of this conditioned response. These results suggest that AMPA receptors are involved in the expression of amphetamine-induced behavioural sensitisation in mice, and that this involvement is limited to either the neurobiological effects of amphetamine or the effects of amphetamine on conditioned associations, rather than drug environment conditioned associations.     

Newborn mice develop balanced Th1/Th2 primary effector responses in vivo but are biased to Th2 secondary responses

Newborn mice are impaired in their abilities to mount protective immune responses. For decades, it was generally held that the poor responses of newborns were largely due to the developmentally immature state of the T cells. In vitro studies showing that neonatal T cells were deficient in Th1 cytokine production, proliferation, and secondary responsiveness strongly supported this idea. Recently, several studies have challenged this view; animals exposed to Ag as neonates were shown to have mature Th1 responses in adulthood. However, it is not clear whether the mature immune responses were actually mounted by T cells generated after the neonatal stage. We have reexamined this issue by analyzing the capabilities of neonatal lymph node T cells to develop into Ag-specific effector cells during the actual neonatal period. Our results demonstrate that the capacity to develop a balanced Th1/Th2 primary effector response is fully mature within the first week of life. However, while neonatal and adult primary cytokine profiles were very similar, Th2 secondary responses predominated in animals first immunized as newborns. Moreover, we have observed other differences between adults and neonatal responses, including 1) the kinetics of cytokine production and responsiveness to adjuvant during the primary response, and 2) the contribution of spleen and lymph node to secondary responses. We propose that these differences reflect developmental regulation of effector cell function that has important consequences to neonatal immune function.     

IL-5 receptor positive B cells, but not eosinophils, are functionally and numerically influenced in mice carrying the X-linked immune defect

Mouse IL-5 (mIL-5) acts on B cells and eosinophils to induce growth and differentiation through the mIL-5 specific receptor (mIL-5R). The functional high-affinity mIL-5R is a heterodimer composed of alpha and beta chains. We investigated the expression of mIL-5R and the responsiveness of B cells and eosinophils to mIL-5 in X-linked immunodeficient (xid) mice. mIL-5R expression analyzed by using mAbs specific for alpha and beta chains revealed that xid B cells had fewer mIL-5R alpha +mIL-5R beta + than BALB/c B cells. In particular, a decrease in the number of peritoneal mIL-5R+ B cells among Ly-1 B cells (known as B-1 cells) was remarkable. Furthermore, the frequency of precursors of mIL-5 responsive B cells in xid mice was approximately 100-fold lower than that of BALB/c mice. Interestingly, sorted mIL-5R+ peritoneal B cells from xid mice displayed a low response to mIL-5. Intraperitoneal injection of mIL-5 into BALB/c mice induced polyclonal IgM production and an increase in the number of eosinophils. The same regimen failed to induce an increase in the same parameters in xid mice. However, xid mice showed mIL-5-induced eosinophilia in peripheral blood to a similar extent as BALB/c mice. Eosinophils from mIL-5-injected xid mice expressed both alpha and beta chains of mIL-5, and responded to mIL-5 with prolonged in vitro survival.     

TCR-gamma delta cells in CD3 zeta-deficient mice contain Fc epsilon RI gamma in the receptor complex but are specifically unresponsive to antigen

Unlike TCR-alpha beta cells, TCR-gamma delta cells express a distinct member of the zeta family, the gamma-chain of Fc epsilon RI (Fc epsilon RI gamma) within the TCR complex. To study the role of the Fc epsilon RI gamma-chain in TCR-gamma delta cells, a TCR-gamma delta transgenic mouse (G8) has been crossed with CD3 zeta-chain-deficient mice (G8.zeta-/-). Thy-1+ spleen and lymph node cells of these animals expressed low levels of CD3/TCR. These results suggested that the zeta-chain is required for effective TCR transport to the cell surface. In contrast, intraepithelial TCR-gamma delta cells of G8.zeta-/- mice expressed high levels of TCR. Immunoprecipitation with anti-CD3 showed that Fc epsilon RI gamma-chains were associated with the TCR complex in T cells isolated from zeta-deficient mice. Although the Fc epsilon RI gamma-expressing T cells proliferated in response to stimulation by TCR-specific Abs including anti-CD3 epsilon, anti-pan gamma delta, and anti-V gamma 2 mAb, the G8.zeta-/- T cells did not respond to the G8-specific Ag (T10b), anti-Thy-1 mAb, or Con A. The unresponsiveness to the Ag was not due to the reduced TCR expression, because intraepithelial TCR-gamma delta cells from the zeta-deficient mice did not respond to Ag. The inability of the G8.zeta-/- T cells to respond to Ag could not be overcome by providing an anti-CD28 costimulatory signal or by adding exogenous rIL-2. Taken together, our data suggest that the Fc epsilon RI gamma-chain associates with the TCR-gamma delta complex in the absence of the zeta-chain, but it is not able to substitute for the zeta-chain for effective transport of TCR to the cell surface or functional responses to Ag.     

Cholesterol and oxygenated cholesterol concentrations are markedly elevated in peripheral tissue but not in brain from mice with the Niemann-Pick type C phenotype

Niemann-Pick disease type C (NP-C) is a rare genetic disorder characterized by progressive neurodegeneration, frequent developmental delay and early death. Tissues of affected individuals accumulate large quantities of free cholesterol in lysosomes. Because cytotoxic oxygenated derivatives of cholesterol are known to form readily when cholesterol concentrations are elevated, we searched for these compounds in liver, kidney, spleen and brain from mice with the NP-C phenotype. In order of abundance, we identified 7 alpha- and 7 beta-hydroxycholesterol, 5 alpha, 6 alpha-epoxycholestan-3 beta-ol, 4 beta-hydroxycholesterol, cholest-4-en-3 beta, 7 alpha-diol and cholest-4-en-3 beta, 6 beta-diol in most tissue samples. Cholesterol concentrations in affected mice were increased 3-fold in kidney and 7- to 8-fold in spleen and liver compared to controls (all p < 0.001) but were unchanged in brain. Although oxysterol levels were markedly elevated in nonbrain tissue, the oxysterol and cholesterol concentrations increased proportionally so that oxysterols expressed as percentage of total sterols were the same for all animals (0.34 +/- 0.19% averaged over all organs in affected animals vs 0.40 +/- 0.42% in control mice). In contrast to peripheral tissue, we could not detect any increase in either absolute or relative oxysterol levels in the brains of affected and control mice (49 +/- 61 vs 53 +/- 43 micrograms/g wet weight and 0.45 +/- 0.52 vs 0.47 +/- 0.37%, respectively). Thus, brain sterols are normal in NP-C mice and it is unlikely that an accumulation of cytotoxic oxygenated derivatives of cholesterol could account for the progressive neuropathology seen in the disease.