Aritake Mizuno

Near-infrared FT-Raman spectra of the rat brain tissues

Near-infrared Fourier transform (FT) Raman spectroscopy was applied to brain tissues in situ. The spectra were obtained from the cerebral cortex, white matter of the cerebrum, caudate-putamen, thalamus, synaptosomal fraction, and myelin fraction. High-quality Raman spectra in the 400 to 2940 cm-1 range were measured without interference of autofluorescence. Common spectral bands were assigned. The ratios of the intensity at 1664 (amide I), 1442 (CH2 deformation), 2885 (CH2 asymmetric stretching), 2938 cm-1 (CH3 symmetric stretching) could be used for differentiation between the gray and white matters.

Biomedical Application of Near-Infrared Fourier Transform Raman Spectroscopy. Part I: The 1064-nm Excited Raman Spectra of Blood and Met Hemoglobin

Raman spectroscopy, which has been employed extensively to study the structure and function of biological materials for the last two decades, also has considerable potential as a diagnostic and analytical tool in medicine. However, the medical application of Raman spectroscopy has been limited to some special topics such as lens research because most morbid materials emit strong fluorescence and are weak with respect to laser illumination. Fluorescence and decomposition, two major drawbacks of Raman spectroscopy which must be circumvented, have made its medical applications difficult.

An application of laser Raman spectroscopy to the study of a hereditary cataractous lens; on the Raman band for a diagnostic marker of cataractous signatures

Raman spectra were measured for cac-strain mouse (Nakano mouse) lenses in the various stages of cataract formation. The spectra were compared with those of normal mouse lenses of the corresponding ages. A significant difference was observed in the intensity of the Raman band due to an OH stretching mode of lens water (3390 cm-1) between the spectra of cataractous lenses and those of normal lenses. The difference was already obvious in the very incipient stage of hereditary cataract and became more pronounced with cataract development. These observations clearly show that changes in lens water occur during cataractogenesis. We propose in this communication that the intensity change of the Raman band at 3390 cm-1 may be very useful for the diagnostic marker of cataractous signatures.

Direct measurement of Raman spectra of intact lens in a whole eyeball

The Raman spectrum of an intact rabbit and rat lens was measured intraocularly without taking it out of a whole eyeball. The Raman spectrum was identical to that of an excised lens. There was no evidence for Raman bands from cornea, aqueous humor, or other ocular tissue in the spectrum of intact lens in the eyeball. The result is an important step toward the clinical application of Raman spectroscopy in the field of cataract research.

Confirmation of lens hydration by Raman spectroscopy

Lens hydration was monitored by laser Raman spectroscopy in WBN/Kob rats which form spontaneous diabetes at about 1 yr of age and develop cataracts approximately 6 months after the onset of diabetes. Lens hydration in response to chronic hyperglycemic stress in these rats appeared predominantly in the cortical region while the hydration state in the nuclear region was fairly preserved. The 9-month treatment of similarly diabetic rats with the aldose reductase inhibitor Ponalrestat sufficiently suppressed lens hydration in the cortical and nuclear regions.

Changes in α-tubulin and actin gene expression during optic nerve regeneration in frog retina

Abstract: The optic nerve of the bullfrog was transected and the regeneration process was investigated. We previously reported that α‐tubulin mRNA in the retina increased to a maximum 1–2 h after optic nerve transection with no specific change in actin mRNA. In the present investigation, we examined the long‐term effect of optic nerve transection. Northern blot analysis revealed that α‐tubulin mRNA increased again gradually after the rapid and transient increase and actin mRNA increased to a maximum at 7 days (more than twofold compared to the control retinas). The period during which actin mRNA reaches a maximal increase almost corresponds to the time lag between the axotomy and the initiation of axonal outgrowth. The main cytoskeletons of neuronal growth cones have been shown to consist of actin containing microfilaments. Therefore, the transient increase of actin mRNA may have a relationship to the initial out growth of axons. On the other hand, the rapid and transient increase of α‐tubulin mRNA observed in our previous studies is probably one of the initial responses of retinal ganglion cells to the axotomy, and the gradual increase in α‐tubulin mRNA observed in this study can probably be interpreted as provision of the structural materials necessary for axonal elongation.

Raman Spectroscopic Study of Cataract Formation: Emory Mouse Cataract

Raman spectra of Emory mouse lenses at various stages of cataract formation have been measured and compared with those of normal lenses from ICR-strain mice. The relative intensity of an OH stretching mode of lens water was considerably stronger for the Emory mouse lenses, even in a precataractous stage, suggesting that lens hydration is deeply implicated in the initiation of Emory mouse cataract. The rate of 2SH → S-S conversion, estimated from the intensity decrease of a SH stretching mode, was similar between the Emory and ICR-strain mouse lenses. Accordingly, it seems unlikely that the conversion is a predominant factor for forming large protein aggregates which lead to lens opacification. A significant change was observed for the intensity ratio of a tyrosine doublet near 840 cm−1 during the cataract development, suggesting that the strength of the hydrogen bonding of some tyrosine residues alters upon the formation of the large protein aggregates. Comparison of the Raman spectral changes observed for Emory mouse cataract with those for normal lens aging and other cataract formation leads us to the conclusion that the microenvironmental change of tyrosine residues is only one common specific feature for lens opacification.

C-Fos mRNA induction under vitamin B6 antagonist-induced seizure

c-Fos mRNA expression was studied in mouse brain after vitamin B6 antagonist-induced seizure. The vitamin B6 antagonists used were hydrazine, thiosemicarbazide, penicillamine and deoxypyridoxine. Only deoxypyridoxine was effective in increasing c-fos mRNA and c-fos protein expression in nerve cells. The other 3 antagonists had levels of c-fos mRNA below or equal to basal level. The seizure activity induced by several vitamin B6 antagonists resulted in different effects on c-fos gene expression.

Effect of L-glutamic acid on [14C]GABA release from isolated rat retina

The effect of various putative neurotransmitters on the release of [14C]GABA taken up by isolated rat retina was investigated by a perfusion technique. L-Glutamic acid initially enhanced the GABA release and subsequently reduced it to a level below the baseline rate. This enhancing effect of L-glutamic acid was Ca2+-dependent and tetrodotoxin-sensitive. L-Aspartic acid only reduced the GABA release, whereas kainic acid exhibited only a marked enhancement of its release. Acetylcholine, norepinephrine, dopamine, glycine and taurine did not have any significant effects on GABA release.

Molecular aging of lens crystallins and the life expectancy of the animal. Age-related protein structural changes studied in situ by Raman spectroscopy

In order to investigate the relationship of molecular aging of lens crystallins to an animals life expectancy or to the type of the lens, Raman spectra have been measured in situ for rabbit and guinea-pig lens nuclei at various stages of aging; these spectra have been compared with those of rat and mouse lens nuclei previously reported. Lens aging results in pronounced differences among the Raman spectra of the lens nuclei of the four species. It is shown that the rates of dehydration, inter- and intramolecular disulfide bond formation, and microenvironmental changes in the tryptophan residues of lens crystallins are different among the four species. Much faster changes occur for rat and mouse, which have a shorter life expectancy (2 years) and give rise to hard lens nuclei while slower changes occur for rabbit and guinea-pig, which have a longer life expectancy (5-7 years), and give soft lens nuclei. In addition, the Raman data reveal, for all the species investigated, that there are correlations among the rates of the dehydration, the inter- and intramolecular disulfide bond formation, and the microenvironmental changes in the tryptophan residues. Therefore, there seems to be a common mechanism for molecular aging of lens crystallins among the four species, although the rate of the molecular aging strongly depends upon the life expectancy of the animal and the type of the lens. The most important factor determining the rate of the molecular aging is probably the dehydration which decreases free water in the lens nucleus.