Tetsuro Horikoshi

RNA molecules were found to separate into numerous metastable conformational forms upon non-denaturing gel electrophoresis. The equilibration of the conformations was accelerated by heating or mild denaturing conditions. Single-base substitutions in the sequence of the RNAs caused changes in the conformational patterns, including mobility shifts of major and minor conformations, appearance of new conformations and loss of other conformations. This sequence-dependent RNA conformational polymorphism was used to detect point mutations in p53 and, dihydrofolate reductase genes. Sense and anti-sense RNA strands corresponding to the same segment of the p53 gene gave entirely different conformational patterns. To generate the RNA, short regions of the target genes (up to about 250 bp) were amplified by the polymerase chain reaction and the resulting DNA segments transcribed to RNA by T7 RNA polymerase. The method is rapid, simple, amenable to non-radioactive visualization and was successful in several cases when DNA single-strand conformational polymorphism analysis (Orita et al. (1989) Genomics 5, 874-879) failed to detect the point mutation.

Lymnaea can be classically conditioned by pairing photic stimulation with a rotational stimulus. The electrophysiological properties of the Lymnaea photoreceptors and statocyst neurons are incompletely known. There are 2 types of ocular photoreceptors and 3 types of statocyst "hair cells." Type A photoreceptors had a response latency from 200 to 400 ms, with a graded depolarizing response having maximum action spectra at 480-500 nm, corresponding to the beta(max) of rhodopsin. Additionally they extend their axons in the direction of the other type of photoreceptor neuron, the type T cell. These neurons have a 2-component response to light: a response reversibly reduced in Ca(2+)-free saline, and a component persisting in Ca(2+)-free saline. Type T cells send processes into the cerebral ganglion and terminate close to the ending of the statocyst hair cells. Hair cells send their terminal branches to the cerebral ganglia close to the terminations of the type T cells. Caudal hair cells respond to a light flash with a depolarization, whereas the rostral cells respond with a hyperpolarization. The response latency in all hair cells was dependent on the stimulus intensity; the brightest light tested had a latency of 200 ms. The photo-induced response was abolished in Ca(2+)-free saline, whereas it was still present in high Ca(2+)-high Mg(2+) saline, consistent with the hypothesis that the connection between the photoreceptors and hair cells is monosynaptic. Thus the sensory information necessary for forming an association between photic and rotational stimuli converges on the statocyst neurons.