Embryonic motoneuron-skeletal muscle co-culture in a defined system.

This paper describes a significant biotechnological advancement by creating a minimalist serum-free defined system to co-culture rat mammalian nerve and muscle cells in order to form functional neuromuscular junctions. To date, all the known in vitro nerve and muscle co-culture models use serum containing media; and while functional neuromuscular junctions (NMJ) are described, they failed to detail or quantify the minimum factors needed to recreate the NMJ in vitro. In this work, we demonstrate the development of a defined motoneuron and muscle co-culture system resulting in the formation of NMJs including: 1) a new culture technique, 2) a novel serum-free medium formulation and 3) a synthetic self-assembled monolayer (SAM) substrate N-1 [3-(trimethoxysilyl) propyl] diethylenetriamine (DETA). We characterized the culture by morphology, immunocytochemistry, electrophysiology and videography. This model system provides a better understanding of the minimal growth factor and substrate interactions necessary for NMJ formation and provides a basic system that can be utilized for nerve-muscle tissue engineering, regenerative medicine and development of limb prosthetics.

Serotonin potentiates the response of neurons of the superficial laminae of the rat spinal dorsal horn to gamma-aminobutyric acid.

Employing the Nystatin-perforated whole-cell patch-clamp recording technique, the modulatory effects of serotonin (5-HT) on gamma-aminobutyric acid (GABA)-activated whole-cell currents were investigated in neurons acutely dissociated from the superficial laminae (laminae I and II) of the rat spinal dorsal horn. The results showed: (1) GABA acted on GABA(A) receptors and elicited inward Cl(-) currents (I(GABA)) at a holding potential (V(H)) of -40 mV; (2) 5-HT potentiated GABA-induced Cl(-) current without affecting the reversal potential of I(GABA) and the apparent affinity of GABA to its receptor; (3) alpha-methyl-5-HT, a selective agonist of 5-HT(2) receptor, mimicked the potentiation effect of 5-HT on I(GABA), whereas ketanserine, an antagonist of 5-HT(2) receptor, blocked the potentiation effect of 5-HT; (4) Chelerythrine, an inhibitor of protein kinase C, reduced the potentiation effect of 5-HT on I(GABA). The present results indicate: (1) The potentiation of 5-HT on I(GABA) is mediated by 5-HT(2) receptor and through a protein kinase-dependent transduction pathway; (2) The interactions between 5-HT and GABA might play an important role in the modulation of nociceptive information transmission at spinal cord level.

Self-assembled monolayers of rigid thiols.

The preparation, structure, properties and applications of self-assembled monolayers (SAMs) of rigid 4-mercapto-biphenyls are briefly reviewed. The rigid character of the biphenyl moiety results in a molecular dipole moment that affects both the adsorption kinetics on gold surfaces, as well as the equilibrium structure of mixed SAMs. Due to repulsive intermolecular interaction, the Langmuir isotherm model does not fit the adsorption kinetics of these biphenyl thiols, and a new Ising model was developed to fit the kinetics data. The equilibrium structures of SAMs and mixed SAMs depend on the polarity of the solution from which they were assembled. Infrared spectroscopy suggests that biphenyl moieties in SAMs on gold have small tilt angles with respect to the surfaces normal. Wetting studies shows that surfaces of these SAMs are stable for months, thus providing stable model surfaces that can be engineered at the molecular level. Such molecular engineering is important for nucleation and growth studies. The morphology of glycine crystals grown on SAM surfaces depends on the structure of the nucleating glycine layer, which, in turn, depends on the H-bonding of these molecules with the SAM surface. Finally, the adhesion of PDMS cross-linked networks to SAM surfaces depends on the concentration of interfacial H-bonding. This non-linear relationship suggests that the polymeric nature of the elastomer results in a collective H-bonding effect.

Isolation of rat sacral dorsal commissural neurons.

UNLABELLED: To isolate rat sacral dorsal commissural neurons (SDCN). METHODS: Using enzymatic and mechanical dissociation techniques to isolate the neurons and using nystatin perforated patch technique to evaluate their functional state. RESULTS: The isolated neurons exhibited good responses to excitatory and inhibitory amino acids. The responses of SDCN to N-methyl-D-aspartate were markedly potentiated by substance P and trans-1-aminocyclopentane-1,3-dicarboxylate, whereas GABA responses were significantly potentiated by diazepam, pregnenolone, and pentobarbital. CONCLUSION: This preparation provides a satisfactory model for exploring the mechanisms of the SDCN in nociception and antinociception.

[Observation on ultrastructure of sterile cysts of Echinococcus granulosus in mice].

The fine structure of sterile cysts of Echinococcus granulosus in mice was studied by TEM. The cyst wall consists of laminated layer and germinal membrane. At ultrastructural level, the laminated layer contains microfibrillate matrix and irregularly shaped electron-dense granules. The germinal membrane comprise of tegument and cell region. Plasmalemma invagination resembling the "pinocytotic vesicle" are seen between the microtriches of the tegument, and mitochondria are seen in the basal portion of the tegument. In the cell region, there are tegumental cells, muscle cells, and glycogen-containing cells. "Nucleolar channel system" can be observed in some of the glycogen-containing cells.

Use of ultrasonography in the diagnosis of dysbaric osteonecrosis.

Comparisons between x-ray and B-mode ultrasonic examinations were carried out in 32 divers in both shoulders and both hips (128 comparisons in all). The x-rays showed dysbaric osteonecrosis (DON) in 46 sites; in 43 of these, the B-mode ultrasonogram result coincided with the x-ray finding. We suggest that B-mode ultrasonograms can depict the location, form, and size of the lesions. It is an easy, efficient, and noninvasive method of diagnosing DON, worthy of further investigation and application. Similar good agreement between x-rays and ultrasonograms was found with the now-obsolete A-mode technique.

Distribution of lesions in the head and neck of the humerus and the femur in dysbaric osteonecrosis.

In a prospective investigation of 164 cases of dysbaric osteonecrosis out of a total of 1269 hyperbaric workers, we found that lesions are apt to occur more frequently in some sites of the head and neck of the femur and humerus than in others. Defining the susceptibility of sites with radiographs has improved our ability to make early diagnoses because we are able to estimate the likelihood of a definite lesion of osteonecrosis developing. We have found that when the trabeculae in a susceptible site appear disordered or have translucent areas, the site often develops into osteonecrosis in a few years.

Ethanol treatment for acute decompression sickness in rabbits.

Rabbits developed acute decompression sickness after staying at 6 ATA for 30 min followed by decompression to 1 ATA in 20 min or less. If the rabbits received an i.v. injection of 25% ethanol upon surfacing, all survived, whereas half the untreated control group died within 15-35 min after decompressing. In ethanol-treated animals, no bubbles were seen in blood vessels of visceral organs, muscles, and subcutaneous tissues at autopsy 60 min after treatment. Decompression reduced platelet counts markedly in all rabbits, but in the control group the count stayed low, whereas with ethanol treatment the counts had reached the precompression level after 1 h and 24 h.