Interrelationship between food availability, fat body, and ovarian cycles in the frog,Rana tigrina, with a discussion on the role of fat body in anuran reproduction

Xavier Santos, Gustavo A. Llorente, Seasonal Variation in Reproductive Traits of the Oviparous Water Snake, Natrix maura, in the Ebro Delta of Northeastern Spain, Journal of Herpetology, Vol. 35, No. 4 (Dec., 2001), pp. 653-660

The amount of body fat is negatively correlated with reproductive activity in Opheodrys aestivus as it is in a number of snakes. Based on experimental evidence from lizards, significant reductions in stored body fat preceding vitellogenesis should adversely affect reproductive output. Comparisons are made between reproductive output in 0. aestivus following a climatically normal year and following an extremely hot and dry year when stored body fat was reduced by approximately 70%. No significant differences were seen in any reproductive parameter between the two years. It is suggested that spring foraging success provided the energy needed for vitellogenesis despite the reduced fat stores. genic activity is important for a number of reptilian species (Derickson, 1976). In lizards lipids may be stored subcutaneously or in ab- dominal fat bodies. However, it is the latter that are the more labile and therefore more im- mediately subject to storage or utilization (Der- ickson, 1974). The amount of lipid in fat bodies approximates the amount in egg clutches (Hahn and Tinkle, 1965). Experimental excision of ab- dominal fat bodies results in a lower rate of yolk deposition, a greater incidence of follicular atresia and delayed or inhibited follicular growth (Hahn and Tinkle, 1965; Smith, 1968). The adaptive significance of fat bodies in lizards ap- pears to be associated with follicular develop- ment during periods of low activity or in early season clutches when feeding conditions are be- low optimum (Smith, 1968; Hahn and Tinkle, 1965). A test of this hypothesis in a natural population of Urosaurus ornatus (Ballinger, 1977) demonstrated that reduced fat storage prior to reproduction resulted in a significant decrease in clutch size and clutch frequency. Low fat storage was attributed to reduced food avail- ability due to low precipitation levels (Ballinger, 1977). Although experimental evidence is lacking, several studies of snakes have shown inverse relationships between vitellogenic activity and fat body weight as occurs in lizards (Volsoe, 1944; St. Girons, 1957; Tinkle, 1962; Wharton, 1966; Gibbons, 1972; Parker and Brown, 1980; Jackson and Franz, 1981; Plummer, in press). Shine (1977) stated that fat was mobilized for follicular development in several species of Aus- tralian snakes. It seems reasonable, therefore, to assume that fat bodies in snakes affect re- production. Cale and Gibbons (1972) showed that fat body weight was a good indicator of total lipid in snakes and suggested that this re- lationship was independent of species. In cen- tral Arkansas female Opheodrys aestivus produce a single clutch annually and typically enter and exit hibernation with about 8% body fat (by weight) stored in the abdominal fat bodies (Plummer, in press). In 1980 after an extremely hot and dry summer, these snakes entered hi- bernation with about 2% body fat. The subse- quent 1981 reproductive output is reported in this paper. METHODS

Anautogeny is a reproductive strategy by which females do not reproduce until they feed. Therefore, nutritional signals must inform the reproductive tissues, and cells that the organism has reached a nutritional status suitable for triggering reproductive processes. One of the possible pathways involved in anautogeny is the "target of rapamycin" (TOR) pathway, which has been described as connecting the nutritional status with growth, proliferation, and cancer. The German cockroach, Blattella germanica, is an anautogenous species whose vitellogenesis is governed by juvenile hormone. In the present report, we describe the cloning of TOR cDNA from B. germanica (BgTOR). Expression studies showed that BgTOR is expressed in adult female corpora allata and fat body. BgTOR knockdown using systemic RNAi in vivo produced a severe inhibition of juvenile hormone synthesis in adult female corpora allata, together with a reduction of mRNA levels corresponding to 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) synthase-1, HMG-CoA synthase-2, and HMG-CoA reductase. In addition, there was a reduction of vitellogenin mRNA in the fat body, and ovaries did not grow. Analysis of TOR expression in corpora allata of fed and starved females suggested that TOR is not regulated at the transcriptional level. Nevertheless, there was a reduction in HMG-CoA synthases and reductase mRNA in corpora allata (but not in the fat body) of starved females, together with a dramatic reduction of juvenile hormone production and ovary development. Taken together, our results indicate that TOR knockdown mimics starvation in terms of corpora allata activity, and suggest that nutritional signals that activate juvenile hormone biosynthesis and vitellogenin production are mediated by the TOR pathway.

The relationship between the annual cycle of energy storage, in the form of abdominal fat bodies, and reproduction was studied in lowland and subalpine populations of the viviparous skink, Ni- veoscincus ocellatus. This species displays an unusual asynchronous reproductive pattern in which male and female preparations for reproduction are separated temporally, thereby providing an opportunity to clarify our understanding about the energy demands of squamate reproduction. Abdominal fat bodies in both sexes showed distinct annual cycles that were positively correlated with resource availability, peaking in summer. In males, fat body mass was correlated positively with testes development and negatively with plasma testosterone concentrations and mating behavior. This is in contrast to the patte obsernved in species with synchronous breeding cycles, where there is generally a negative relationship between testis size and lipid storage. We concluded that N. ocellatus is able to produce spermatozoa and develop abdominal fat reserves concurrently, and therefore testis size is unlikely to be constrained by energetic costs. In contrast, mating behavior appears to rely on stored energy, even though this results in reduced fat reserves for winter hi- bernation. Female fat bodies were smallest in midspring, coinciding with the end of vitellogenesis and early pregnancy. Vitellogenesis and early pregnancy appear to be energetically expensive and are reliant on stored energy in this species. Large fat bodies at the end of pregnancy in both populations are surprising given the reported high costs of reproduction. Two factors may contribute to this: resource availability may be greater in late summer, and the energetic costs associated with pregnancy appear to be highest in the early to midstages of gestation.

One way that aerobic biological systems counteract the generation of reactive oxygen species (ROS) is with superoxide dismutase proteins SOD1 and SOD2 that metabolize superoxide radicals to molecular oxygen and hydrogen peroxide or scavenge oxygen radicals produced by the extensive oxidation-reduction and electron-transport reactions that occur in mitochondria. We characterized SOD1 and SOD2 of Bombyx mori isolated from the fat body of larvae. Immunological analysis demonstrated the presence of BmSOD1 and BmSOD2 in the silk gland, midgut, fat body, Malpighian tubules, testis and ovary from larvae to adults. We found that BmSOD2 had a unique expression pattern in the fat body through the fifth instar larval developmental stage. The anti-oxidative functions of BmSOD1 and BmSOD2 were assessed by exposing larvae to insecticide rotenone or vasodilator isosorbide dinitrate, which is an ROS generator in BmN4 cells; however, exposure to these compounds had no effect on the expression levels of either BmSOD protein. Next, we investigated the physiological role of BmSOD1 and BmSOD2 under environmental oxidative stress, applied through whole-body UV irradiation and assayed using quantitative RT-PCR, immunoblotting and microarray analysis. The mRNA expression level of both BmSOD1 and BmSOD2 was markedly increased but protein expression level was increased only slightly. To examine the differences in mRNA and protein level due to UV irradiation intensity, we performed microarray analysis. Gene set enrichment analysis revealed that genes in the insulin signaling pathway and PPAR signaling pathway were significantly up-regulated after 6 and 12 hours of UV irradiation. Taken together, the activities of BmSOD1 and BmSOD2 may be related to the response to UV irradiation stress in B. mori. These results suggest that BmSOD1 and BmSOD2 modulate environmental oxidative stress in the cell and have a specific role in fat body of B. mori during pupation.

The free abdominal fat body of adult female Mediterranean field crickets, Gryllus bimaculatus, synthesizes lipids from [1-(14)C]-acetate in vitro. Up to an age of 12h, the incorporation of acetate into lipids is very low and then rises to a maximum 24h after adult emergence. Thereafter, the incorporation of acetate decreases to moderate levels at day 2 and then slowly decreases until day 30. The adipokinetic hormone of G. bimaculatus (Grb-AKH) significantly inhibits the incorporation of acetate at a concentration of 10(-11) M; maximum inhibition (approximately 95%) is reached at 10(-8) M. The inhibiting effect of Grb-AKH is fast, dose-dependent, and reversible. The periovaric fat body shows a similar pattern of acetate incorporation, although rates of incorporation are lower; the incorporation can be inhibited by Grb-AKH as well. The segmental abdominal fat body and the fat body from the head both incorporate acetate into lipids at low rates that cannot be inhibited significantly by AKH. Prepurified brain extracts significantly inhibit acetate incorporation by free abdominal fat bodies at a concentration of 0.1 brain equivalent. Allatostatins and crustacean cardioactive peptide, which are both present in cricket brains, are not responsible for this inhibiting effect. Octopamine causes a dose-dependent inhibition of acetate incorporation whereas synephrine had no such effect. The inhibiting effect of Grb-AKH on the formation of lipid stores in the fat body and its consequences for reproductive processes are discussed.

Ovarian follicular dynamics and fluctuations in fat body, oviducal, and liver masses were studied in captive Rana cyanophlyctis in comparison with wild-caught frogs, sampled at monthly intervals over a period of 12 months. In both the captive and wild-caught frogs first growth phase (FGP) and second growth phase (SGP) or vitellogenic oocytes were produced throughout the period examined; however, changes in ovarian and oviducal weights were less marked in the former group. In the captive frogs SGP oocyte production was reduced by 50%, and, maximum ovarian weight and SGP oocyte number were attained 2-3 months earlier than in wild-caught controls. The FGP oocyte pool in laboratory-maintained frogs, however, was comparable with that of the corresponding wild-caught frogs. Captivity caused a threefold increase in atresia and reduced the number of oocytes reaching SGP. The depletion of fat stores in fat bodies during the later phases of captivity suggests that the deposition of lipids into oocytes (for SGP) was given priority over storage in the fat bodies. The low oviducal weights of captive frogs was correlated with a reduced number of SGP oocytes, which are the source of estrogen. On the other hand, liver weight remained high, indicating adequate hepatic vitellogenin synthesis. Possible reduction in its output was not detected, possibly due to the reduced number of follicles reaching SGP. The findings indicate that stress of captivity decreases gonadotrophins and estrogen levels. Oviducal growth is reduced in captive frogs.(ABSTRACT TRUNCATED AT 250 WORDS)

M. boulengeri is oviparous; males are in breeding condition in spring. H, peronii is viviparous; males are in breeding condition in autumn and winter. The former has abdominal fat bodies; the latter does not. The amounts of fat in tails, fat bodies, eggs, and the rest of the body were measured at different times of the year in both species. Changes in these fat stores suggest that fat from tails and, where present, fat bodies, is used by females for producing eggs and by males for activities associated with reproductive activity. In both sexes of both species there were large differences between individuals in the amount of stored fat. Removing tails from females made them produce fewer eggs; the reduction was greater in H, peronii. When M. boulengeri were fasting, they used more fat from their tails than from any other source, but they used up a higher proportion of fat body fat than of their other fat stores.

Role of Temperature and Photoperiod in the Onset of Sexual Maturity in Female Frogs, Rana cyanophlyctis

In reptiles, many lipid reserve structures were recognized, and different patterns of storage and utilization of lipids have been identified. In this study, a population of Liolaemus koslowskyi was studied to analyze the diversity and functioning of the lipid reserve structures. The specific objectives were to identify the main lipid reserve structures, and to describe their seasonal patterns; to study sexual and seasonal variations of the reserve structures; and to analyze the relation between seasonal patterns of such reserves with the sexual cycle and other biological aspects. Individuals of both sexes collected at five different times of the year were analyzed. The abdominal fat bodies, livers, tails, fat deposits of the lateral folds of the neck, the fat bodies of the axillary region and gonads were weighted. The fat content of livers, tails and neck fat reserves were determined through the extraction in a Soxhlet device. The volume of the ovaries, oviductal oocytes and testicles, and the average surface and thickness of the epididymides were estimated. The abdominal fat bodies and the tail-extracted fats were the main lipid reserves. In general, females showed higher lipid contents, and significant seasonal variations in weight and lipid content in both sex were found, while seasonal variations in the liver fresh weight and liver fat content were found only in males. The axillar and neck fats deposits probably function as auxiliary reserves, and the caudal autotomy did not significantly affect tail fat content. The sexual cycles were synchronic in both sexes.

Role of fat body and uterine gland in milk synthesis by adult female Glossina morsitans

Aedes vigilax mosquitoes, collected from the south coast of New South Wales, were fed on a blood-meal containing the T48 strain of Ross River virus. Viral infection and multiplication in this major vector were studied, together with dissemination of the virus in organs and tissues as determined by immunofluorescence. Individuals of this field mosquito population were highly susceptible to infection (ID50 = 10(3.4) BHK-21 TCID50 per mosquito) and virus replicated rapidly, reaching a maximum level 5-6 days after feeding. Specific viral antigens were detected in sections of abdominal midgut, abdominal fat bodies, thoracic fat bodies, cerebral ganglia and head fat bodies 2-4 days after ingestion of the virus-containing bloodmeal. Antigens were also detected 5-7 days after feeding in abdominal ganglia, thoracic ganglia, salivary glands and foregut. Antigens were demonstrated in sections of eggs from the ovary of one mosquito, which provided further evidence of transovarial transmission of Ross River virus in Ae. vigilax.

Fatty acid distribution of lipids from carcass, liver and fat bodies of the lizard, Cnemidophorus tigris, prior to hibernation

Glycolytic and lipolytic effects of ovine FSH and estradiol-17β in the lizard Anolis carolinensis

Most animals can modulate nutrient storage pathways according to changing environmental conditions, but in honey bees nutrient storage is also modulated according to changing behavioral tasks within a colony. Specifically, bees involved in brood care (nurses) have higher lipid stores in their abdominal fat bodies than forager bees. Pheromone communication plays an important role in regulating honey bee behavior and physiology. In particular, queen mandibular pheromone (QMP) slows the transition from nursing to foraging. We tested the effects of QMP exposure on starvation resistance, lipid storage, and gene expression in the fat bodies of worker bees. We found that indeed QMP-treated bees survived much longer compared to control bees when starved and also had higher lipid levels. Expression of vitellogenin RNA, which encodes a yolk protein that is found at higher levels in nurses than foragers, was also higher in the fat bodies of QMP-treated bees. No differences were observed in expression of genes involved in insulin signaling pathways, which are associated with nutrient storage and metabolism in a variety of species; thus, other mechanisms may be involved in increasing the lipid stores. These studies demonstrate that pheromone exposure can modify nutrient storage pathways and fat body gene expression in honey bees and suggest that chemical communication and social interactions play an important role in altering metabolic pathways.