Light reduces the excitation efficiency in the nss mutant of the sheep blowfly Lucilia

The nss (no steady state) phototransduction mutant of the sheep blowfly Lucilia was studied electrophysiologically using intracellular recordings. The effects of the nss mutation on the receptor potential are manifested in the following features of the light response. (a) The responses to a flash or to dim lights are close to normal, but the receptor potential decays close to the baseline level during prolonged illumination after a critical level of light intensity is reached. (b) The decline of the response is accompanied by a large reduction in responsiveness to light that recovers within 20 s in the dark. (c) The full reduction in responsiveness to light is reached when approximately 13% of the photopigment molecules are converted from rhodopsin (R) to metarhodopsin (M). (d) A maximal net pigment conversion from R to M by blue light induces persistent inactivation in the dark, without an apparent voltage response. This inactivation could be abolished at any time by M-to-R conversion with orange light. The above features of the mutant indicate that the effect of the nss mutation on the light response of Lucilia is very similar to the effects of the transient receptor potential (trp) mutation on the photoreceptor potential of Drosophila. Noise analysis and voltage measurements indicate that the decay of the receptor potential is due to a severe reduction in the rate of occurrence of the elementary voltage responses (bumps). The bumps are only slightly modified in shape and amplitude during the decline of the response to light of medium intensity. There is also a large increase in response latency during intense background illumination. These results are consistent with the hypothesis that separate, independent mechanisms determine bump triggering and bump shape and amplitude. The nss mutation affects the triggering mechanism of the bump.     

Ectopic expression of β-lactoglobulin/human serum albumin fusion genes in transgenic mice: hormonal regulation andin situ localization

We produced transgenic mice carrying the native sheep -lactoglobulin (BLG) or fusion genes composed of the BLG promoter and human serum albumin (HSA) minigenes. BLG was expressed exclusively in the mammary glands of the virgin and lactating transgenic mice evaluated. In contrast, transgenic females carrying the BLG/HSA fusion constructs also expressed the HSA RNA ectopically in skeletal muscle, kidney, brain, spleen, salivary gland and skin. Ectopic expression of HSA RNA was detected only in strains that express the transgene in the mammary gland. There was no obvious correlation between the level of the HSA RNA expressed in the mammary gland and that found ectopically. In three transgenic strains analysed, the expression of HSA RNA in kidney and skeletal muscle increased during pregnancy and lactation, whereas in the brain HSA expression decreased during lactation in one of the strains. HSA protein was synthesized in skeletal muscle and skin of strain #23 and its level was higher in lactating mice compared with virgin mice. Expression of HSA was also analysed in males and was found to be more stringently controlled than in females of the same strains.In situ hybridization analyses localized the expressed transgene in the skin, kidney, brain and salivary glands of various transgenic strains. Distinct strain-specific and cell-type specific HSA expression patterns were observed in the skin. This is in contrast to the exclusive expression of the HSA transgene in epithelial cells surrounding the alveoli of the mammary gland. Taken together, these results suggest that the absence of sufficient mammary-specific regulatory elements in the BLG promoter sequences and/or the juxtaposition of the BLG promoter with the HSA coding sequences leads to novel tissue- and cell-specific expression in ectopic tissues of transgenic mice.     

Regulation of glycogen synthetase and phosphorylase phosphatase activities in rat adipose tissue.

Incubation of a rat adipose tissue homogenate causes a time and temperature dependent activation of glycogen synthetase (UDP glucose:glycogen 4-alpha-glucosyltransferase) and simultaneous inactivation of phosphorylase (1,4-alpha-D-glucan: orthophosphate alpha-glucosyltransferase, EC 2.4.1.1). Activation of glycogen synthetase at 15 and 23 degrees C was preceded by a lag period. The duration of the lag period could not be correlated with significant changes in phosphorylase activity. Addition of glucose and methylxanthines caused an increase in the rates of glycogen synthetase activation and phosphorylase inactivation. The effect on glycogen synthetase activation was mainly on the linear phase. Addition of AMP inhibited phosphorylase inactivation and accelerated glycogen synthetase activation. Addition of muscle phosphorylase alpha caused a prolongation of the lag period which lasted until phosphorylase alpha activity had decreased to the level originally present in the preparation. It is concluded that in adipose tissue activation of glycogen synthetase is not dependent on prior inactivation of phosphorylase and that other factors should be looked for to explain the lag period preceding glycogen synthetase activation.     

Kinetic and immunologic evidence for the absence of glucose-6-phosphatase in early human chorionic villi and term placenta.

The existence of the enzyme glucose-6-phosphatase (G6Pase) in early and term human placenta was investigated by comparing the characteristics of placental microsomal glucose 6-phosphate (G6P) hydrolytic activity and liver G6Pase. Placental microsomes exhibited similar apparent Km values for G6P and beta-glycerophosphate in intact and deoxycholate-treated microsomes, heat stability at acidic pH, low latency of mannose 6-phosphate hydrolysis, very low activity of pyrophosphate: glucose phosphotransferase, and undetectable [U-14C]G6P transport into the placental microsomes, all of which indicated that specific G6Pase activity does not exist in placenta. Immunological evidence of the absence of both 36.5 kDa and T2 proteins, which represent the G6Pase catalytic protein and the phosphate/pyrophosphate transporter protein, respectively, confirmed that early and term human placenta are devoid of the multicomponent G6Pase enzyme.     

Isozymes of glutamate dehydrogenase from oat leaves: Properties and light effect on synthesis

Ammonium-dependent induction of a GDH isozyme in oat leaveswas proportional to light intensity and inhibited by DCMU. Thestimulation of GDH synthesis in response to ammonia was partiallyrepressed by benzimidazole. The inducible (no. 1) and noninducible(no. 2) GDH isozymes wereseparated and purified by approximately54 and 24 fold respectively. The two isozymes were highly specificfor NAD and the rate of NADH oxidation was 7 to 9 times higherthan NAD reduction. Both isozymes showed similar Km values forsubstrates of the reductive amination reaction and pH optimafor NADH oxidation. The pH optima for NAD reduction were 9 and8.2 for isozymes1 and 2 respectively. The two isozymes had asimilar molecular weight, 2.2–2.4 x 10⁵ but differed intheir isoelectric point and temperature sensitivity. Resultssuggest that the GDH isozymes in oat leaves are two differententities but might possess a similar metabolic function. (Received January 6, 1976; )