Cobalt and nickel coordination polymers containing 3-pyridylnicotinamide and five-membered ring dicarboxylates

<div><p>Cobalt and nickel coordination polymers containing the conformationally flexible 3-pyridylnicotinamide (3-pna) ligand and a five-membered ring-based dicarboxylate ligand have been prepared and structurally characterized via single-crystal X-ray diffraction. [Co(tpdc)(3-pna)]_n (<b>1</b>) was prepared using 2,5-thiophenedicarboxylic acid (H₂tpdc). This material shows a 2-D layer structure containing {Co₂(OCO)₂} dimers linked by tpdc and 3-pna ligands. Compound <b>1</b> manifests an underlying 3,5-connected (4²6)(4²6⁷8) 3,5L2 topology. The isostructural pair of solids [Co(<i>D</i>-cam)(3-pna)(H₂O)₂]_n (<b>2</b>) and [Ni(<i>D</i>-cam)(3-pna)(H₂O)₂]_n (<b>3</b>) was obtained from the chiral <i>D</i>-camphorate (<i>D</i>-cam) ligand. These two materials possess acentric ribbon coordination polymer motifs. Compound <b>1</b> manifests antiferromagnetic coupling concomitant with Kramers doublet formation. Thermal properties of these materials are also discussed.</p></div

Structural analysis of NMJs from 4-6 week-old BALB/c and <i>nmf375</i> mice.

<p><i>A</i>, The NMJs of the tibialis anterior (TA) and triceps brachii (TRI) muscles from <i>nmf375</i> and BALB/c mice were stained for neurofilament (green) and α-bungarotoxin (red) and imaged using confocal microscopy. Left: 20X magnification, scale bar 50 μm; Right: 63X magnification, scale bar 20 μm. <i>B</i>, Comparison of NMJs from TA muscles of <i>ax</i>^<i>J</i> and <i>nmf375</i> mice stained as in A. Note the NMJs from <i>ax</i>^<i>J</i> muscle contain terminal swellings (white arrows).</p

Hippocampal paired pulse facilitation (PPF) in BALB/c, <i>nmf375</i>, C57, and <i>ax</i>^<i>J</i> mice at 4-5 weeks of age.

<p>Dendritic field EPSPs were measured in the CA1 region of the hippocampus in response to extracellular stimulation of Schaffer collateral axons. Two stimuli, spaced at the time intervals indicated, were given at 0.1Hz, and the paired pulse ratio at each interval was obtained by dividing the initial slope of the second pulse by the initial slope of the first pulse (n= 3-4 animals per group).</p

Analysis of <i>nmf375</i> and <i>ax</i>^<i>J</i> mutations on C57BL/6J and BALB/c backgrounds.

<p><i>A</i>, Survival curves for wt (C57BL/6J and BALB/c), BALB<i>ax</i>^J and C57<i>nmf375</i>. N=4 mice per genotype. <i>B</i>, The trypsin-like activity of the 20S proteasome in arbitrary fluorescence units (AFU) from BALB/c and C57 mice was examined at 4 and 8 weeks of age (n = 2-4 per genotype, per age; p = 0.15 and 0.80, respectively). <i>C</i>, Western blot of proteasomal subunits in whole brain and proteasomal fractions from BALB/c and C57 mice (n = 2-3 per genotype). <i>D</i>, Representative western blot of monomeric ubiquitin in whole brain extracts from embryonic day 17-E18 C57, C57<i>ax</i>^<i>J</i>, and C57<i>nmf375</i> mice (n = 3 per genotype). <i>E</i>, Quantitation of monomeric ubiquitin levels (2-way ANOVA; *p-value < 0.05). Data were analyzed by Student’s t-test and are shown as mean ±SEM.</p

Analysis of NMJ structure, ubiquitin levels, and muscle AChR subunit expression in 10-12-week-old BALB/c and <i>nmf375</i> mice.

<p><i>A</i>, Confocal images of whole-mount immunostaining of TA muscles stained with antibodies for neurofilament (green) and α-bungarotoxin (red). Terminals from <i>nmf375</i> mutants displayed accumulations of neurofilaments not observed in controls (white arrows). 63X magnification, scale bar 50 μm. <i>B</i>-<i>C</i>, Quantitation of the area (<i>B</i>) and size distribution (<i>C</i>) of the motor endplates. <i>D</i>, Quantitation of AChR subunit expression in gastrocnemius muscle (***p-value <0.001, n = 4). <i>E</i>, Immunoblot of ubiquitin from spinal cord extracts of BALB/c and <i>nmf375</i> mice. Quantitation of the immunoblot revealed a 55% reduction in monoubiquitin (**p < 0.01, n = 3). <i>F</i>, Representative immunoblot of total brain extracts from BALB/c and <i>nmf375</i> mice probed for polyubiquitin conjugates. <i>G</i>, Analysis of forelimb grip strength. <i>H</i>, Rotarod analysis <b><i>I</i></b>, von Frey analysis of tactile sensitivity. For <b><i>G-I</i></b>, n = 4-6 mice per genotype. *p-value <0.05, **p-value <0.01, ***p-value <0.001. Data were analyzed by Student’s t-test and are shown as mean ±SEM.</p

Examination of muscle AChR subunit expression and fiber type cross-sectional area of gastrocnemius muscle from 4-6 week-old BALB/c and <i>nmf375</i> mice.

<p><i>A</i>, qPCR of α, β, δ, γ, and ε AChR subunit mRNAs from gastrocnemius muscles; *p-value <0.05; **p-value <0.01, ***p-value <0.001. n = 3 per genotype. <i>B</i>-<i>C</i>, Quantitation of type I (B) and type II (C) fiber cross-sectional area from gastrocnemius muscles (n = 3 per genotype, 25-60 fibers per animal). Data were analyzed by Student’s t-test and are shown as mean ±SEM.</p

Characterization of <i>Usp14</i> in the <i>nmf375</i> mice.

<p><i>A</i>, Schematic diagram of <i>Usp14</i> cDNA species amplified. Open box indicates exonic sequence, lower case letters denote intronic sequence, grey box indicates intronic sequence maintained in mature transcript, angled lines represent splicing pattern. <i>B</i>, Genomic DNA sequence of <i>Usp14</i> exon 9/intron 9 border from <i>nmf375</i>, BALB/c, C57BL/6J, C3H/HeJ, FVB, and DBA mice. Capital letters denote exonic sequence; lower case letters and grey box denotes intronic sequence. Asterisk and dark grey box denotes mutation. <i>C</i>, Representative qPCR of <i>Usp14</i>, <i>Uch37, Poh1</i> mRNA from spinal cord extracts of 4-6 week old BALB/c and <i>nmf375</i> mice (n = 3 per genotype). Data were analyzed by Student’s t-test and are shown as mean ±SEM. <i>D</i>, Representative immunoblot of USP14 from spinal cord extracts from 4-6 week old BALB/c and <i>nmf375</i> mice (n = 3 per genotype). Blots were also probed for β-tubulin as a loading control.</p

<i>Usp14</i> maps to the <i>nmf375</i> critical region.

<p><i>A</i>, Meiotic and physical linkage map of the <i>nmf375</i> critical region. Filled circle denotes the centromere, open box represents critical region. <i>B</i>, Survival curves of wt (C57BL/6J and BALB/c), <i>ax</i>^<i>J</i>, and <i>nmf375</i> mice. <i>C</i>, Body weights from 4- to 20-week-old C57BL/6J, BALB/c, <i>ax</i>^<i>J</i>, and <i>nmf375</i> mice. <i>D</i>, Open field Analysis. <b><i>E</i></b>, Measurements of forelimb grip strength. <i>F</i>, Rotarod Analysis. <b><i>G</i></b>, Von Frey assay for tactile sensitivity. <i>D</i>-<i>G</i>, mice were 4-6 weeks of age. *p-value <0.05, **p-value <0.01, ***p-value <0.001. (n = 4-6 mice per genotype). Data were analyzed by Student’s t-test and are shown as mean ±SEM.</p

Analysis of ubiquitin expression in <i>nmf375</i> mutants at 4-6 weeks of age.

<p><i>A</i>, Representative immunoblot of monomeric ubiquitin from spinal cord extracts BALB/c and <i>nmf375</i> mice (n = 4). <i>B</i>, Representative immunoblot of polyubiquitin conjugates from BALB/c and <i>nmf375</i> spinal cord extracts. <i>C</i>, Quantitation of monomeric ubiquitin levels from A (p = 0.12). <i>D</i>, Representative qPCR of ubiquitin <i>Ubb</i> and <i>Ubc</i> mRNA from spinal cord extracts of BALB/c and <i>nmf375</i> mice (n=3 per genotype); *p-value <0.01. <i>E</i>, Active DUBs in proteasome extracts isolated from the brains of BALB/c and <i>nmf375</i> mice were labeled with HA-UB-VME and probed with an anti-HA antibody. Blots were also probed for the α1 subunit (RPT1) of the 19S proteasome as a loading control (n = 3 per genotype). <i>F</i>, Representative immunoblot of proteasome extracts from BALB/c and <i>nmf375</i> mice probed for POH1, UCH37, USP14 and RPT1. Data were analyzed by Student’s t-test and are shown as mean ±SEM.</p

Effect of tau reduction on development and survival of <i>ax^J</i> mice.

<p>(A) Four-month survival curves of wt, <i>ax^J</i>, <i>tau^KO</i> and <i>ax^Jtau^KO</i> mice. <i>n</i> = 4 mice per genotype. (B) Body weights of 4 and 8-week-old wt, <i>ax^J</i>, <i>tau^KO</i> and <i>ax^Jtau^KO</i> mice. <i>n</i> = 4 mice per genotype. (C) Purkinje cell axonal swellings were quantitated in wt, <i>ax^J</i>, <i>tau^KO</i> and <i>ax^Jtau^KO</i> mice. n = 4 mice per genotype. ***indicates p≤10⁻⁷ (D) Paired-pulse facilitation was measured at hippocampal CA3-CA1 synapses. Facilitation was measured at 5 interpulse intervals ranging from 50 to 500 ms. n = 4 mice per genotype.</p