Extended fear conditioning reveals a role for both N-methyl-D-aspartic acid and non-N-methyl-D-aspartic acid receptors in the amygdala in the acquisition of conditioned fear

Pavlovian conditioning is a useful tool for elucidating the neural mechanisms involved with learning and memory, especially in regard to the stimuli associated with aversive events. The amygdala has been repeatedly implicated as playing a significant role in the acquisition and expression of fear. If the amygdala is critical for the acquisition of fear, then it should contribute to this processes regardless of the parameters used to induce or evaluate conditioned fear. A series of experiments using reversible inactivation techniques evaluated the role of the amygdala in the acquisition of conditioned fear when training was conducted over several days in rats. Fear-potentiated startle was used to evaluate the acquisition of conditioned fear. Pretraining infusions of N-methyl-d-aspartic acid (NMDA) or non-NMDA receptor antagonists alone into the amygdala interfered with the acquisition of fear early in training, but not later. Pretraining infusions of a cocktail consisting of both an NMDA and non-NMDA antagonist interfered with the acquisition of conditioned fear across all days of training. Taken together these results suggest the amygdala may potentially be critical for the acquisition of conditioned fear regardless of the parameters utilized.     

Virostatic action of rimantidin studied on a phage-bacterium model

The action of rimantidin on reproduction of phages T3, T4, and was investigated. The maximal inhibitory action of the compound was observed between the first and sixth minute of infection with phage T3, but no such effect was observed on replication of phages T4 and . It is postulated on the basis of the results of incorporation of labeled precursors into phage DNA, RNA, and protein and the character of manifestation of the action of the compound on the phage yield that the inhibitory effect of rimantidin is evidently determined by its effect on bacterial and phage RNA polymerases, mainly on the latter.Research Laboratory of Experimental Immunobiology, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR N. N. Zhukov-Verezhnikov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 81, No. 5, pp. 564–566, May, 1970.     

The study of biogenetic pathways using a perfusion technique containing perfluorochemicals.

A liver perfusion system was assembled and adapted for pulse labelling studies. The perfusion medium was prepared by emulsifying perfluorotributylamine and Pluronic F 68 in a CO2 atmosphere using a sonicator. Ribosome-rich and ribosome-poor rough microsomes, smooth microsomes and Golgi membranes could be prepared from perfused livers with a good purity and recovery as from non-perfused livers. The subfractionation technique used was simple and involved slight modifications of the methods described earlier by Eriksson (1973) and Ehrenreich et al. (1973). The specific activity of NADPH-cytochrome c reductase in microsomes and of UDP-galactosyltransferase in Golgi membranes from perfused and non-perfused livers were identical. The specific activity of glucose-6-phosphatase in microsomes was slightly decreased after perfusion, but the membrane permeability barrier to glucose-6-phosphate remained intact. The granulated microsomal fractions from perfused liver had a somewhat reduced number of membrane-bound ribosomes. The system developed proved useful in studies of the synthesis and intracellular transport of albumin. The technique should also be suitable for use in studies of membrane biogenesis.     

Porous titanium scaffolds fabricated using a rapid prototyping and powder metallurgy technique

One of the main issues in orthopaedic implant design is the fabrication of scaffolds that closely mimic the biomechanical properties of the surrounding bone. This research reports on a multi-stage rapid prototyping technique that was successfully developed to produce porous titanium scaffolds with fully interconnected pore networks and reproducible porosity and pore size. The scaffolds' porous characteristics were governed by a sacrificial wax template, fabricated using a commercial 3D-printer. Powder metallurgy processes were employed to generate the titanium scaffolds by filling around the wax template with titanium slurry. In the attempt to optimise the powder metallurgy technique, variations in slurry concentration, compaction pressure and sintering temperature were investigated. By altering the wax design template, pore sizes ranging from 200 to 400 microm were achieved. Scaffolds with porosities of 66.8 +/- 3.6% revealed compression strengths of 104.4+/-22.5 MPa in the axial direction and 23.5 +/- 9.6 MPa in the transverse direction demonstrating their anisotropic nature. Scaffold topography was characterised using scanning electron microscopy and microcomputed tomography. Three-dimensional reconstruction enabled the main architectural parameters such as pore size, interconnecting porosity, level of anisotropy and level of structural disorder to be determined. The titanium scaffolds were compared to their intended designs, as governed by their sacrificial wax templates. Although discrepancies in architectural parameters existed between the intended and the actual scaffolds, overall the results indicate that the porous titanium scaffolds have the properties to be potentially employed in orthopaedic applications.     

Human heart beat analysis using a modified algorithm of detrended fluctuation analysis based on empirical mode decomposition

How to quantify the complexity of a physiological signal is a crucial issue for verifying the underlying mechanism of a physiological system. The original algorithm of detrended fluctuation analysis (DFA) quantifies the complexity of signals using the DFA scaling exponent. However, the DFA scaling exponent is suitable only for an integrated time series but not the original signal. Moreover, the method of least squares line is a simple detrending operation. Thus, the analysis results of the original DFA are not sufficient to verify the underlying mechanism of physiological signals. In this study, we apply an innovative timescale-adaptive algorithm of empirical mode decomposition (EMD) as the detrending operation for the modified DFA algorithm. We also propose a two-parameter scale of randomness for DFA to replace the DFA scaling exponent. Finally, we apply this modified algorithm to the database of human heartbeat interval from Physiobank, and it performs well in identifying characteristics of heartbeat interval caused by the effects of aging and of illness.     

A new class of agonists and antagonists of N-methyl-D-aspartic acid receptors: Derivatives of imidazole-4,5- and pyrazole-3,4-dicarboxylic acids

Studies of imidazole-4,5- and pyrazole-3,4-dicarboxylic acid derivatives revealed a number of new agonists and antagonists of N-methyl-D-aspartic acid (NMDA) receptors. Studies were based on whole-cell patch-clamp methods applied to rat hippocampus pyramidal cells. Increases in the lipophilicity of the environment of the nitrogen atom, keeping the distance between the terminal acid functions constant, led to a weakening of NMDA antagonism and increases in NMDA antagonism. Increases in the lipophilicity around the nitrogen atom could also lead to less of selectivity in the interaction with NMDA receptors and the appearance of non-NMDA antagonist properties. Translated from Rossiiskii Fiziologicheskii Zhurnal imeni I. M. Sechenova, Vol. 85, No. 4, pp. 523–530, April, 1999.     

6,7-Dichloro-3-hydroxy-2-quinoxalinecarboxylic acid is a relatively potent antagonist at NMDA and kainate receptors

The 6,7-dichloro derivative of 3-hydroxy-2-quinoxalinecarboxylic acid (diCl-HQC) is a relatively potent antagonist at the two excitatory amino acid receptor subtypes activated by N-methyl-D-aspartate (NMDA) and kainic acid. It antagonizes NMDA-induced excitation in the frog spinal cord (pA2 5.8), i.e. with a potency similar to D-2-amino-7-phosphonoheptanoate (D-AP-7). It also antagonizes NMDA-induced sodium efflux from rat brain slices (pA2 5.6). The compound inhibits kainic acid-induced sodium efflux from rat brain slices with a pA2 of 5.4 and it inhibits [3H]kainic acid binding to rat brain membranes with a pKi of 5.4. DiCl-HQC is only weakly active at the quisqualate receptor. This spectrum of activities may make this compound a useful tool to investigate the pharmacology of excitatory amino acid receptors.     

Greater contribution of N-methyl-D-aspartic acid receptors in ventral compared to dorsal hippocampal slices in the expression and long-term maintenance of epileptiform activity

Functional segregation along the dorso-ventral axis of the hippocampus is a developing concept. The higher susceptibility of the ventral hippocampus to epileptic activity compared with dorsal hippocampus is one of the main features, which still has obscure mechanisms. Using the model of magnesium-free medium and field recordings, single epileptiform discharges displayed higher incidence (77% vs 57%), rate (41.7+/-3.1 vs 13.5+/-0.7 events/min), duration (173.9+/-17.7 vs 116.8+/-13.6 ms) and intensity (coastline, 25.4+/-2.5 vs 9.5+/-1.8) in ventral compared with dorsal rat hippocampal slices. In addition, the decay phase of the evoked synaptic potentials was 110% slower in ventral slices. The N-methyl-D-aspartate (NMDA) receptor antagonist d-(-)-2-amino-5-phosphonopentanoic acid (50-100 microM) decreased the discharge rate and coastline similarly in ventral and dorsal slices, but it shortened the discharges in ventral slices (by 40%) only. The NMDA receptor antagonist 3-((R)-2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (10 microM) decreased the rate in both groups and additionally shortened discharges in both kinds of slices, an effect which was greater in ventral ones (31% vs 13%). Furthermore, both drugs shortened the evoked potentials more in ventral (77%) than in dorsal slices (52%). On the other hand, 1 microM of 3-((R)-2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid shortened the discharges and evoked synaptic potentials only in ventral slices, and slowed down the discharge rate only in dorsal slices. Addition of NMDA, in the magnesium-free medium, enhanced activity in both kinds of slices. At 5 and 10 microM of NMDA 51% of the ventral but only 9% of the dorsal slices displayed persistent epileptiform discharges, which were recorded for at least one hour after reintroduction of magnesium in the medium. At 10-20 microM the enhancement of activity was transient, followed by suppression of discharges in 40% and 76% of the ventral and dorsal slices, respectively. Most of the slices having experienced suppression did not develop persistent activity. We propose that the NMDA receptors contribute to the higher susceptibility of the ventral hippocampus to expression and long-term maintenance of epileptiform discharges. This diversification may be related to other aspects of hippocampal dorso-ventral functional segregation.     

The technique of gait analysis using a Coda-3 motion analysis system

Gait analysis using Code-3 provides information which allows individual gait to be characterised with little interference to the patient. It is an ideal method for analysing gait in patients with cerebral palsy. Because of the ease with which this information is now available using Coda-3, gait analysis can now be performed on all ambulant patients in whom surgery is contemplated.     

Sleep-promoting action of excitatory amino acid antagonists: a different role for thalamic NMDA and non-NMDA receptors

Changes in the sleep-waking cycle of freely moving cats were studied during application of excitatory amino acid antagonists in the ventro-posterolateral thalamic nuclei by microdialysis. DL-2-Amino-5-phosphono-pentanoic acid (APV), a selective N-methyl-D-aspartate (NMDA) receptor antagonist, produced an increase in the deep stages of slow wave sleep and in paradoxical sleep and a decrease in the light stages of slow wave sleep (SWS1), while 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), at a concentration selective for the non-NMDA receptors, produced a marked increase in SWS1. These results indicate a strong sleep-promoting action of excitatory amino acid antagonists and suggest that thalamic NMDA and non-NMDA receptors may play different roles in sleep regulation. Thus, changes in the sleep-waking cycle should be carefully evaluated when assessing the potential clinical use of excitatory amino acid antagonists.     

Participation of nitric oxide and N-methyl-D-aspartic acid receptors in the pressor response to intrathecal injected noradrenaline at the spinal cord of the rat

The upregulated expression of tetrodotoxin sensitive (TTXs) Na+ channels is thought to play an important role in the development of ectopic discharges (EDs) in axotomized sensory neurons. The present study examined the levels of mRNAs of three subtypes of TTXs Na(+) channels, Na(v)1.7, Na(v)1.6, and Na(x), in the dorsal root ganglion (DRG) after segmental spinal nerve ligation. Following nerve ligation, the level of mRNAs of Na(v)1.7 and Na(v)1.6 was decreased, while the Nax mRNA level was increased at 5 days, but not at 1 day, postoperatively compared with the normal levels. Thus, if upregulated expression of TTXs Na+ channels contributes to the generation of EDs in axotomized DRG neurons, Na(x) is the most likely contributor among the three tested subtypes.     

A rapid and efficient microtechnique for the analysis of functional transferrin receptors on tumor cells

A simple and efficient method for the analysis of the affinity and number of functional transferrin receptors (TFR) on human tumor cells is described. The technique is designed to utilize microtitration equipment; and is suitable for easy comparison of up to 8 different cell preparations per assay. Using this technique, 5 established cell lines were evaluated for functional TFR expression. The control erythroleukemic cell line K562 possessed 3.28 X 10(5) functional TFR per cell (+/- 3.69 X 10(4), SEM) Kd = 9.0 X 10(-9) X M-1. Trypsin and heat-pretreated cells were compared to control erythroleukemic K562 cells from the same culture to determine both the effects of receptor removal and cell viability on the assay. Trypsin and heat pretreatment of these K562 cells severely decreased receptor function as indicated by Scatchard analysis as well as by time course and cold competition analysis respectively. Whereas the affinity of trypsin-treated receptors on cells was similar to control values, heat-killed cells displayed an altered cellular affinity for 125I-transferrin underscoring the importance of utilizing cells of high viability in receptor assays.     

Error analysis of the quantification of hepatic perfusion using a dual-input single-compartment model

We performed an error analysis of the quantification of liver perfusion from dynamic contrast-enhanced computed tomography (DCE-CT) data using a dual-input single-compartment model for various disease severities, based on computer simulations. In the simulations, the time-density curves (TDCs) in the liver were generated from an actually measured arterial input function using a theoretical equation describing the kinetic behavior of the contrast agent (CA) in the liver. The rate constants for the transfer of CA from the hepatic artery to the liver (K(1a)), from the portal vein to the liver (K(1p)), and from the liver to the plasma (k(2)) were estimated from simulated TDCs with various plasma volumes (V(0)s). To investigate the effect of the shapes of input functions, the original arterial and portal-venous input functions were stretched in the time direction by factors of 2, 3 and 4 (stretching factors). The above parameters were estimated with the linear least-squares (LLSQ) and nonlinear least-squares (NLSQ) methods, and the root mean square errors (RMSEs) between the true and estimated values were calculated. Sensitivity and identifiability analyses were also performed. The RMSE of V(0) was the smallest, followed by those of K(1a), k(2) and K(1p) in an increasing order. The RMSEs of K(1a), K(1p) and k(2) increased with increasing V(0), while that of V(0) tended to decrease. The stretching factor also affected parameter estimation in both methods. The LLSQ method estimated the above parameters faster and with smaller variations than the NLSQ method. Sensitivity analysis showed that the magnitude of the sensitivity function of V(0) was the greatest, followed by those of K(1a), K(1p) and k(2) in a decreasing order, while the variance of V(0) obtained from the covariance matrices was the smallest, followed by those of K(1a), K(1p) and k(2) in an increasing order. The magnitude of the sensitivity function and the variance increased and decreased, respectively, with increasing disease severity and decreased and increased, respectively, with increasing stretching factor except for V(0). Identifiability analysis showed that the identifiability between K(1)(p) and k(2) was lower than that between K(1)(a) and k(2) or between K(1a) and K(1p). In conclusion, this study will be useful for understanding the accuracy and reliability of the quantitative measurement of liver perfusion using a dual-input single-compartment model and DCE-CT data.     

A rapid solution-phase screening technique for hybridoma culture supernatants using radiolabeled antigen and a solid-phase immunoadsorbent

The solid-phase immunoassay commonly used to screen hybridoma supernatants at times gave falsely positive results. A solution-phase screening technique, which uses radiolabeled antigen and a solid-phase immunoadsorbent, is described. This technique overcomes the problem of false positives and can be easily adopted for other soluble antigens.