The North American Free Trade Agreement: Reasons For Passage And Requirements To Be A Foreign Legal Consultant In A Nafta Country

Under a global economy, countries around the world trade with each other. International trade is essential to achieving a global economy. As the world moves toward a global economy, the need for international trade of goods and services is increasing. Numerous agreements between countries have been passed to facilitate international trad

Survival of adult generated hippocampal neurons is altered in circadian arrhythmic mice.

The subgranular zone of the hippocampal formation gives rise to new neurons that populate the dentate gyrus throughout life. Cells in the hippocampus exhibit rhythmic clock gene expression and the circadian clock is known to regulate the cycle of cell division in other areas of the body. These facts suggest that the circadian clock may regulate adult neurogenesis in the hippocampus as well. In the present study, neurogenesis in the hippocampal subgranular zone was examined in arrhythmic Bmal1 knockout (-KO) mice and their rhythmic heterozygous and wildtype littermates. Proliferation and survival of newly generated subgranular zone cells were examined using bromodeoxyuridine labelling, while pyknosis (a measure of cell death) and hippocampal volume were examined in cresyl violet stained sections. There was no significant difference in cellular proliferation between any of the groups, yet survival of proliferating cells, 6 weeks after the bromodeoxyuridine injection, was significantly greater in the BMAL1-KO animals. The number of pyknotic cells was significantly decreased in Bmal1-KO animals, yet hippocampal volume remained the same across genotypes. These findings suggest that while a functional circadian clock is not necessary for normal proliferation of neuronal precursor cells, the normal pruning of newly generated neurons in the hippocampus may require a functional circadian clock

The volume of the hippocampus does not differ between wildtype, heterozygous and <i>Bmal1-</i>KO mice.

<p>A) Cavalieri estimates of total hippocampal volume in <i>Bmal1</i> (+/+), (+/−), and (−/−) mice at approximately 100 days of age. Estimates were calculated from the cresyl violet stained tissue series utilized in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0099527#pone-0099527-g003" target="_blank">Figure 3</a> for pyknotic cell counts. Hippocampal volume estimates were not significantly different across genotypes.</p

<i>Bmal1</i>-KO exhibit decreased pyknosis, indicative of diminished cell death, in the dentate gyrus relative to heterozygous and wildtype mice.

<p>A) Photomicrograph of cresyl violet stained pyknotic cells, shown by the white arrows. Scale bar = 50 µm. B) Estimates of the total number of GCL pyknotic cells throughout the extent of the hippocampi of <i>Bmal1</i> (+/+), (+/−), and (−/−) mice, calculated by counting pyknotic cells in every fourth slice and multiplying by 4. *<i>p</i><0.05.</p

<i>Bmal1</i>-KO exhibit enhanced survival of newly generated cells in the dentate gyrus relative to heterozygous and wildtype mice.

<p>A) Representative photomicrographs of BrdU (green) and NeuN (red) immunoreactivity in <i>Bmal1</i> (+/+), (+/−), and (−/−) mice 6 weeks after an i.p. injection of BrdU. Scale bar = 200 µm. B) High-magnification confocal image of NeuN/BrdU labeled cells. Scale bar = 10 µm. C) Total number of BrdU labelled cells throughout the hippocampus, calculated by counting cells in every fourth slice and multiplying by 4. *<i>p</i><0.05.</p

Homozygous <i>Bmal1</i>-KO mice are arrhythmic.

<p>Representative wheel-running activity charts (actograms) from <i>Bmal1</i> (+/+), (+/−) and (−/−) animals. Every horizontal line represents 2 days of activity, with subsequent days plotted to the right of and below the previous day. Vertical deflections from the horizontal represent 10 minute bins in which wheel revolutions were detected, with the height of the deflection being proportional to the number of revolutions. The animals were housed in a light/dark cycle for the first 4 weeks (darkness indicated by grey shading) and were then transferred into constant darkness for another 4 weeks. Wildtype (left) and heterozygous (middle) animals had normal circadian patterns of activity in both the light/dark cycle as well as in constant darkness. The <i>Bmal1</i>-KO mice were arrhythmic throughout. Animals in this circadian activity assessment were different than those used in the subsequent proliferation/survival experiments.</p

<i>Bmal1</i>-KO do not exhibit proliferation differences in the sub granular zone of the dentate gyrus.

<p>A) Photomicrographs of DAB labelled BrdU positive cells in <i>Bmal1</i> (+/+), (+/−) and (−/−) SGZ of the DG, 24 hours after an i.p. injection of BrdU. Scale bar = 200 µm. B) Number of cells labelled with BrdU throughout the extent of the hippocampus, calculated by counting cells in every fourth slice and multiplying by 4. Differences between genotypes were not significant.</p