Electrophysiological properties of isthmic neurons in frogs revealed by in vitro and in vivo studies

The frog nucleus isthmi (parabigeminal nucleus in mammals) is a visually responsive, cholinergic and anatomically well-defined group of neurons in the midbrain. It shares reciprocal topographic projections with the ipsilateral optic tectum (superior colliculus in mammals) and strongly influences visual processing. Anatomical and biochemical information indicates the existence of distinct neural populations within the frog nucleus isthmi, which raises the question: are there electrophysiological distinctions between neurons that are putatively classified by their anatomical and biochemical properties? To address this question, we measured frog nucleus isthmi neuron cellular properties in vitro and visual response properties in vivo. No evidence for distinct electrophysiological classes of neurons was found. We thus conclude that, despite the anatomical and biochemical differences, the cells of the frog nucleus isthmi respond homogeneously to both current injections and simple visual stimuli.     

A fourth complementation group among ionizing radiation-sensitive Chinese hamster cell mutants defective in DNA double-strand break repair.

The XR-V9B mutant of Chinese hamster V79 cells which exhibits hypersensitivity to ionizing radiation was isolated by the replica plating technique. The increased sensitivity of XR-V9B cells to X rays (approximately 4-fold, as judged by the D10) was accompanied by increased sensitivity to other DNA-damaging agents such as bleomycin (approximately 17-fold), VP16 (approximately 6-fold), and adriamycin (approximately 5-fold). Only a slightly increased sensitivity was observed after exposure to UV radiation, MMS, or mitomycin C (1.4-, 1.7-, and 2-fold, respectively). As measured by neutral elution after exposure to X rays, XR-V9B cells showed a defect in the rejoining of double-strand breaks (DSBs); after 4 h of repair more than 50% of DSBs remained in comparison to 5% in wild-type cells. No difference was observed in the kinetics of single-strand break rejoining between XR-V9B and wild-type cells, as measured by alkaline elution. To determine whether XR-V9B represents a new complementation group among ionizing radiation-sensitive Chinese hamster cell mutants defective in DSB repair, XR-V9B cells were fused with XR-V15B, XR-1, and V-3 cells, which have impaired DSB rejoining and belong to three different complementation groups. In all cases, the derived hybrids regained the sensitivity of wild-type cells when exposed to X rays, indicating that the XR-V9B mutant represents a new fourth complementation group among X-ray-sensitive Chinese hamster cell mutants defective in DSB repair.     

Recent advances in ABE fermentation: hyper-butanol producing Clostridium beijerinckii BA101

This is an overview of the mutant strain Clostridium beijerinckii BA101 which produces solvents (acetone–butanol–ethanol, ABE) at elevated levels. This organism expresses high levels of amylases when grown on starch. C. beijerinckii BA101 hydrolyzes starch effectively and produces solvent in the concentration range of 27–29 g l⁻¹. C. beijerinckii BA101 has been characterized for both substrate and butanol inhibition. Supplementing the fermentation medium (MP2) with sodium acetate enhances solvent production to 33 g l⁻¹. The results of studies utilizing commercial fermentation medium and pilot plant-scale reactors are consistent with the results using small-scale reactors. Pervaporation, a technique to recover solvents, has been applied to fed-batch reactors containing C. beijerinckii BA101, and solvent production as high as 165 g l⁻¹ has been achieved. Immobilization of C. beijerinckii BA101 by adsorption and use in a continuous reactor resulted in reactor productivity of 15.8 g l⁻¹ h⁻¹. Recent economic studies employing C. beijerinckii BA101 suggested that butanol can be produced at US$0.20–0.25 lb⁻¹ by employing batch fermentation and distillative recovery. Application of new technologies such as pervaporation, fed-batch culture, and immobilized cell reactors is expected to further reduce these prices. Journal of Industrial Microbiology & Biotechnology (2001) 27, 287–291. Received 12 September 2000/ Accepted in revised form 27 January 2001     

CAG repeat analyses in frozen and formalin-fixed tissues following primer extension preamplification for evaluation of mitotic instability of expanded SCA1 alleles

Neurodegenerative disorders, including spinocerebellar ataxias (SCA), Huntington disease (HD) and dentatorubral-pallidoluysian atrophy (DRPLA), are associated with unstable CAG repeats. To investigate the mitotic stability of the repetitive element in the genes for SCA1, SCA3, HD, and DRPLA we extracted DNA from up to 13 tissue samples from four deceased individuals with progressive neurological disorders and neuropathological signs. Due to the formalin fixation of some tissues the genomic DNA was highly degraded and unsuitable for amplification of fragments longer than 150 bp. After size selection and primer extension preamplification, specific analyses could be performed even for expanded alleles. In all four patients the SCA1 mutation could be demonstrated, in one case with remarkable somatic heterogeneity of the elongated allele, whereas alleles of the normal range were stable in all tissues examined. Received: 3 February 1997 / Accepted: 9 April 1997     

Roles for focal adhesion kinase (FAK) in blastomere abscission and vesicle trafficking during cleavage in the sea urchin embryo

Is focal adhesion kinase (FAK) needed for embryonic cleavage? We find that FAK is expressed during early cleavage divisions of sea urchin embryos as determined by polyclonal antibodies to the Lytechinus variegatus protein. FAK is absent in eggs and zygotes and then cycles in abundance during the first cleavages after fertilization. It is maximal at anaphase, similar to the destruction and synthesis of cyclin proteins. To investigate whether FAK is needed during early cleavage, we interfered with its function by microinjecting eggs with anti-FAK antibodies or with FAK antisense morpholino oligonucleotides. Both treatments led to regression of the cleavage furrow. FAK knockdown with antibodies or morpholino oligonucleotides also resulted in an over-accumulation of endocytic vesicles. Thus, FAK could be restricting endocytosis or increasing exocytosis in localized areas important for abscission. FAK appears to be necessary for successful cleavage. These results are the first to document a functional role for FAK during embryonic cleavage.