Single 5 µm diameter needle electrode block modules for unit recordings in vivo.

Investigations into mechanisms in various cortical areas can be greatly improved and supported by stable recording of single neuronal activity. In this study, fine silicon wire electrodes (diameter 3 µm, length 160 µm) are fabricated by vapor-liquid-solid (VLS) growth with the aim of stabilizing recording and reducing the invasiveness on the measurement procedure. The electrode is fabricated on a modular 1 × 1 mm2 conductive silicon block that can be assembled into a number of different device packages, for example on rigid or flexible printed circuit boards (PCB). After plating with a 5 µm diameter platinum black, the needle exhibits an electrical impedance of ~100 kΩ at 1 kHz in saline. The in vivo recording capability of the device is demonstrated using mice, and spike signals with peak-to-peak amplitudes of 200-300 µV in the range 0.5-3 kHz are stably detected, including single-unit activities in cortical layer 2/3. In addition, the device packaged with a flexible PCB shows stable unit recordings for 98.5 min (n = 4). Consequently, our modular, low-invasive needle electrode block devices present an effective route for single-unit recordings in vivo, as well as demonstrating adaptability in device design for a diverse range of experiments.

Resetting central and peripheral circadian oscillators in transgenic rats.

In multicellular organisms, circadian oscillators are organized into multitissue systems which function as biological clocks that regulate the activities of the organism in relation to environmental cycles and provide an internal temporal framework. To investigate the organization of a mammalian circadian system, we constructed a transgenic rat line in which luciferase is rhythmically expressed under the control of the mouse Per1 promoter. Light emission from cultured suprachiasmatic nuclei (SCN) of these rats was invariably and robustly rhythmic and persisted for up to 32 days in vitro. Liver, lung, and skeletal muscle also expressed circadian rhythms, which damped after two to seven cycles in vitro. In response to advances and delays of the environmental light cycle, the circadian rhythm of light emission from the SCN shifted more rapidly than did the rhythm of locomotor behavior or the rhythms in peripheral tissues. We hypothesize that a self-sustained circadian pacemaker in the SCN entrains circadian oscillators in the periphery to maintain adaptive phase control, which is temporarily lost following large, abrupt shifts in the environmental light cycle.

Oxidation of lipids in low density lipoprotein particles.

This study was undertaken to understand further the mechanisms and dynamics of the oxidation of lipids in low density lipoprotein (LDL) particles, aiming specifically at elucidating the material balance between oxygen uptake and products found and also the relative susceptibilities to oxidation of cholesteryl ester in the core and phosphatidylcholine in the outer monolayer in the LDL particles. It was found that considerable amount of oxygen uptake could not be accounted for by conjugated diene or total peroxides. Total peroxide was measured from the phosphine oxide formed from triphenylphosphine or diphenyl-pyrenylphosphine by reduction of peroxides. Cholesteryl ester hydroperoxides and phosphatidylcholine hydroperoxides were the major peroxides formed in LDL oxidation, but they accounted for about 60% of total peroxide. Cholesterol was also oxidized, but its oxidation was significant only at the later stages of the reaction. It was also found that the oxidizability of cholesteryl ester relative to phosphatidylcholine was larger within the LDL particle than in homogeneous solution and this was interpreted in the context of the physical properties of LDL particle.

2,2'-Azobis (4-methoxy-2,4-dimethylvaleronitrile), a new lipid-soluble azo initiator: application to oxidations of lipids and low-density lipoprotein in solution and in aqueous dispersions.

Both hydrophilic and hydrophobic azo radical initiators are useful for in vitro studies on lipid peroxidation and its inhibition by antioxidants. In the present study, a new lipophilic azo compound, 2,2'-azobis(4-methoxy-2,4-dimethylvaleronitrile) (MeO-AMVN), was introduced and its action as an initiator of lipid peroxidation was examined. MeO-AMVN decomposed about 15 times as fast as 2,2'-azobis(2,4-dimethylvaleronitrile) (AMVN), a widely used lipophilic azo initiator, and MeO-AMVN-initiated free radical-mediated peroxidations of lipids in organic solution and in micelles, membranes, and low-density lipoprotein in aqueous dispersions with much smaller concentration than AMVN. The rate of chain initiation by MeO-AMVN varied significantly with the medium and decreased with increasing viscosity of the medium. The advantage and cautions for using MeO-AMVN as a lipophilic radical source have been discussed and it has been concluded that MeO-AMVN, when properly used, is a useful radical initiator of lipid peroxidations especially in micelles, membranes, and lipoproteins.

Identification of the mammalian homologues of the Drosophila timeless gene, Timeless1.

We have identified novel mammalian homologues of a Drosophila clock gene, timeless, and designated them as human TIMELESS1 (hTIM1) and mouse Timeless1 (mTim1), respectively. These genes were mapped by FISH to chromosomal regions 12q12-13 in human and 10D3 in mouse. The deduced amino acid sequences of hTim1 and mTim1 proteins were 1208 and 1197 amino acids in length and shared 83% identity. Northern blot analysis identified a single transcript of 4.5 kb expressed widely in many tissues examined. Unlike the Drosophila counterpart, the levels of the mTim1 transcript exhibited no prominent circadian oscillation in the mouse brain.