Purification of cone visual pigments from chicken retina

A novel method for purification of chicken cone visual pigments was established by use of a 3-[(3-cholamidopropyl)dimethylammonio]-1- propanesulfonate-phosphatidylcholine (CHAPS-PC) mixture. Outer segment membranes isolated from chicken retinas were extracted with 0.75% CHAPS supplemented with 1.0 mg/mL phosphatidylcholine (CHAPS-PC system). After the extract was diluted to 0.6% CHAPS, it was loaded on a concanavalin A-Sepharose column. Elution from the column with different concentrations of methyl alpha-mannoside yielded three fractions: the first was composed of chicken violet, blue, and red in roughly equal amounts, the second predominantly contained chicken red, and the third was rhodopsin with a small amount of chicken green, which was separated from rhodopsin by DEAE-Sepharose column chromatography. Since CHAPS has little absorbance at both ultraviolet and visible regions, we could demonstrate the absolute absorption spectra of chicken red (92%) and rhodopsin (greater than 96%) in these regions. The maximum of the difference spectrum between either chicken red or rhodopsin and its photoproduct (all-trans-retinal oxime plus opsin) was determined to be 571 or 503 nm, respectively. Although chicken green was contaminated with a small amount of rhodopsin having a similar spectral shape, the maximum of its difference spectrum was located at 508 nm by taking advantage of the difference in susceptibility against hydroxylamine between these pigments. Although chicken blue and chicken violet were minor pigments present in the first fraction from the concanavalin A column, their maxima in the difference spectra were determined to be at 455 and 425 nm, respectively, by a partial bleaching method.(ABSTRACT TRUNCATED AT 250 WORDS)     

Multiple visual pigments in a photoreceptor of the salamander retina

Although a given retina typically contains several visual pigments, each formed from a retinal chromophore bound to a specific opsin protein, single photoreceptor cells have been thought to express only one type of opsin. This design maximizes a cell''s sensitivity to a particular wavelength band and facilitates wavelength discrimination in retinas that process color. We report electrophysiological evidence that the ultraviolet-sensitive cone of salamander violates this rule. This cell contains three different functional opsins. The three opsins could combine with the two different chromophores present in salamander retina to form six visual pigments. Whereas rods and other cones of salamander use both chromophores, they appear to express only one type of opsin per cell. In visual pigment absorption spectra, the bandwidth at half-maximal sensitivity increases as the pigment''s wavelength maximum decreases. However, the bandwidth of the UV-absorbing pigment deviates from this trend; it is narrow like that of a red-absorbing pigment. In addition, the UV-absorbing pigment has a high apparent photosensitivity when compared with that of red- and blue-absorbing pigments and rhodopsin. These properties suggest that the mechanisms responsible for spectrally tuning visual pigments separate two absorption bands as the wavelength of maximal sensitivity shifts from UV to long wavelengths.     

Gonadotropin secretion and testicular function in golden hamsters exposed to skeleton photoperiods with ultrashort light pulses

Both sexually mature and sexually regressed male golden hamsters were transferred to asymmetric skeleton photoperiods with night interruptions of varying duration, the short pulses occurring 14 h after "dawn." Testicular function and accompanying changes in follicle-stimulating hormone (FSH), luteinizing hormone (LH), testosterone and spermatogenesis were observed. Sexually regressed animals exposed to a night-break of 6 seconds (sec) or longer exhibited maximal testicular development with a rapid rise in FSH secretion followed by a slower, more variable rise in LH. Full testicular size was achieved after 8 weeks. Night-breaks of 250 milliseconds (msec) or 1 sec induced testicular development and spermatogenesis but at a slower rate: levels of FSH and LH were still rising at the end of the experiment. Complete testicular maintenance was achieved by night-breaks of 1 sec or longer. Partial testicular regression was observed with a night-break of 250 msec. Night-breaks (60 sec) given less frequently than daily also stimulated testicular function and a night-break every 7 days increased FSH and LH secretion in sexually regressed hamsters, causing testicular development at a submaximal rate. Night-breaks given more frequently induced rapid testicular growth. Almost complete testicular maintenance of sexually mature hamsters was achieved with a 60-sec night-break at weekly intervals. Symmetric skeleton photoperiods also triggered testicular development in sexually regressed hamsters, with two 1-sec light pulses (14 h apart) being almost as effective as a normal long day. No difference in reproductive function was observed between animals on long days (14L:10D) and those exposed to maximally stimulatory skeleton photoperiods.     

Microenvironmental regulation of visual pigment expression in the chick retina

Visual pigment (VP) expression in the chick embryo retina was investigated in ovo, in dissociated and explant cultures, and in cDNAs from individual cells. While VP mRNA is not detectable by in situ hybridization until embryonic day (ED) 14-16 in ovo, analysis of VP expression by RT-PCR showed that VP messages are present in the retina as many as 7-10 days before they become detectable by in situ hybridization, and are also detected in other regions of the embryonic CNS. On the other hand, red opsin expression is markedly accelerated when cells are isolated from their intraocular microenvironment at ED 6, and placed in pigment epithelium-free dissociated or explant cultures. This acceleration occurs regardless of cell density, birth date, or serum presence in the medium, suggesting that many photoreceptors are already programmed to express red opsin on or before ED 6, and that microenvironmental inhibitory factors prevent implementation of this program until ED 14 in ovo. The selectivity of this phenomenon is suggested by the finding that other VPs are not observed by in situ hybridization in ED 6 cultures, although they are detectable in cultures of older retinas. Taken together, these findings suggest that red opsin expression may be constitutive for many developing photoreceptor cells in the chick.     

Twenty-four-hour profiles of prolactin and testosterone in ram lambs exposed to skeleton photoperiods consisting of various light pulses

Individual groups of 6 ram lambs were housed within a controlled environment and exposed to one of 6 photoperiod schedules. Groups I and II received 8 (short day) or 16 (long day) h of continuous light, respectively; Groups III, IV and V were exposed to asymmetrical skeleton photoperiods consisting of a main light period of 7 h followed 9 h later by a light pulse of 1 h, 15 min or 1 min duration, respectively, and Group VI was exposed to a symmetrical skeleton photoperiod consisting of two 1-h light pulses positioned 16 h apart. After 4 weeks of treatment serum concentrations of prolactin and testosterone were measured over 24 h. Long-day responses characteristic of the 16L:8D photoperiod (i.e. elevated prolactin and reduced testosterone) were obtained in each of the asymmetric light-pulse treatment groups, but whereas prolactin was elevated over the full 24 h in lambs exposed to 16L:8D, two prominent nocturnal prolactin releases were largely responsible for the high 24-h mean prolactin values in Groups III, IV and V. Reduced serum testosterone in these same groups could not be attributed to a diurnal pattern of secretion but was associated with an overall decrease in testosterone pulse frequency. Prolactin and testosterone levels in Group IV were intermediate between those observed in lambs exposed to 8 or 16 h of light. In summary, light pulses of short duration (1 min) positioned at 17 h after dawn can produce endocrine changes in lambs similar to those observed in lambs exposed to 16 h of continuous light.     

Polyphosphoinositide hydrolysis in response to light stimulation of rat and chick retina and retinal rod outer segments

Phosphoinositides of chick and rat retina were labelled with [3H]inositol. Exposure of retinal preparations to light for 30 s caused loss of labelled phosphatidylinositol 4,5-bisphosphate and to a smaller extent of the other phosphoinositides. Similar light-induced changes were seen when rod outer segment preparations were used and, when these were illuminated in calcium-free media, phosphatidylinositol 4,5-bisphosphate was the only lipid affected. No inositol 1,4,5-trisphosphate was seen after either 30 s or 5 s of illumination of retina or 30 s illumination of rod outer segments. It is concluded that this compound plays no direct part in vertebrate photoreceptor light transduction, though phosphoinositide metabolism might relate to adaptation mechanisms.     

On the three visual pigments in the retina of the firefly squid,Watasenia scintillans

Summary The deep-sea bioluminescent squid, Watasenia scintillans, has three visual pigments: The major one (A1 pigment) is based on retinal and has _max = 484 nm, the second one (A2 pigment) is based on 3-dehydroretinal and has _max = 500 nm, and the third one (A4 pigment) is based on 4-hydroxyretinal and has _max = 470 nm. The distribution of these 3 visual pigments in the retina was studied by HPLC analysis of the retinals in retina slices obtained by microdissection. It was found that A1 pigment was not located in the specific region of the ventral retina receiving the down-welling light which contains very long photoreceptor cells, forming two strata. A2 and A4 pigment were found exclusively in the proximal pinkish stratum and in the distal yellowish stratum. The role of these pigments in the retina is hypothesized to involve spectral discrimination. The extraction and analysis of retinoids to determine the origin of 3-dehydroretinal and 4-hydroxyretinal in the mature squid showed only a trace amount of 4-hydroxyretinol in the eggs. Similar analysis of other cephalopods collected near Japan showed the absence of A2 or A4 pigment in their eyes.Abbreviations HPLC high-performance liquid chromatography - IS inner segment - OS outer segment     

Fly visual pigments difference in visual pigments of blowfly and dronefly peripheral retinula cells

Journal of Comparative Physiology A - The visual pigments of peripheral retinula cells in fly eyes have been investigated by microspectrophotometry in vivo. Since flies have a pupil mechanism...     

Changes in redox states of respiratory pigments recorded from the eyes of live blowflies exposed to light stimuli and hypoxia

Time courses of mitochondrial responses to illumination-induced physiological loads and to hypoxia, were recorded optically from eyes of blowflies Calliphora vicina chalky. We isolated changes in redox states of haems a₃, a, c, and b. Two types of responses to light stimulation were observed. Haems b and a₃ responded with transient oxidation and haems a and c with reduction. The same two groups emerged in response to anoxic exposure. The onset of reduction of haems a and c had virtually no latency, while haems a₃ and b exhibited a transient oxidation followed by reduction only after 10–20 s. The dependence of the steady-state reduction level on P_textO2 P_{{{text{O}}_{2} }} produced the same groups. Haems a and c were significantly reduced at P_textO2 P_{{{text{O}}_{2} }} levels around 10 kPa while with haems b and a₃ load-induced oxidation was only replaced by reduction below 2 kPa. We propose haems respond to physiological loads in accordance with their steady-state reduction, which in turn depends largely on barriers for electron transport imposed by the mitochondrial membrane potential. We also propose it may be possible to assess the values of tissue P_textO2 P_{{{text{O}}_{2} }} and O₂ consumption by monitoring haems that are highly oxidized at rest such as haem a.     

Culture of chick embryo neural retina in serum-free medium : Expression of cholinergic proteins

Dissociated retinal cells from 8-day-old chick embryos were cultivated in serum-containing and in defined serum-free media. Under the latter conditions, and using polylysine as a substrate, the proliferation of glial cells was almost completely prevented, and pure (>90%) neuronal cultures could be maintained for up to 7 days in vitro. The specific but not the total activities of choline acetyltransferase and of the nicotinic and the muscarinic acetylcholine receptor were increased under serum-free culture conditions. Autoradiographic studies with [¹²⁵I]α-bungarotoxin, a selective ligand for nicotinic cholinergic receptors, showed that serum-free culture conditions may constitute a useful tool for identifying biochemically different types of retinal neurons in tissue culture.     

Evolution of visual pigments in geckos

Most geckos are nocturnal forms and possess rod retinas, but some diurnal genera have pure-cone retinas. We isolated cDNAs encoding the diurnal gecko opsins, dg1 and dg2, similar to nocturnal gecko P521 and P467, respectively. Despite the large morphological differences between the diurnal and nocturnal gecko photoreceptor types, they express phylogenetically closely related opsins. These results provide molecular evidence for the reverse transmutation, that is, rods of an ancestral nocturnal gecko have backed into cones of diurnal geckos. The amino acid substitution rates of dgl and dg2 are higher than those of P521 and P467, respectively. Changes of behavior regarding photic environment may have contributed to acceleration of amino acid substitutions in the diurnal gecko opsins.     

Gonadotrophin secretion and pituitary responsiveness to LHRH in castrated and intact male rabbits exposed to different photoperiods

Adult male wild rabbits were exposed to at least 16 weeks of 16L:8 D before experiments began. Plasma LH and FSH concentrations increased significantly (P less than 0.001) when rabbits were castrated in 16L:8D but declined when rabbits were transferred to 8L:16D. Concentrations had returned to normal for castrated rabbits in 16L:8D by 74 days after the start of the 8L:16D treatment. Treatment of intact male rabbits with an injection of LHRH before and after transfer to short daylengths caused a transient increase in plasma LH which lasted 50-80 min and this produced a concomitant rise in plasma testosterone. The daylength change had no effect on this response even though testicular size declined after the transfer to short daylengths. Rabbits moulted in response to exposure to 8L:16D. This suggests that hypothalamic activity responds to photoperiod and that changes in pituitary responsiveness to LHRH and steroid negative feedback are unimportant.