pregnancy requires an increase in insulin secretion - induced glucose metabolism , in order to compensate for the insulin resistance at the end of gestation . in pregnant rats , the threshold of glucose - stimulatedinsulinsecretionbegins to diverge from controls by day 10 and islet cell proliferation and insulinsecretory profiles are inhibited by day 20 to avoid gestational diabetes .
return to normal values ofinsulinsecretionand inhibition ofcelldivision is particularly important when the adequate nutrient intake is not provided during pregnancy .
for example , maternal low - protein diet ( 6% casein ) causes a loss of glucose sensitivity and secretory capacity in pancreatic islets , which is probably the result of alterations to the coupling of stimuli with insulin secretion .
in fact , a maternal low - protein diet decreases insulin secretion and impairs glucose homeostasis , leading to gestational diabetes mellitus and increased risk of chronic diseases in later life [ 1 , 4 ] .  
recently , it has been recognized that an active maternal lifestyle , including regular to moderate physical activity , improves aerobic fitness and the maternal - fetal physiological reserve by enhancing nutrient and oxygen availability to the fetus [ 57 ] .
maternal physical activity is associated with normalised body weight gain , enhanced cardiorespiratory , and improved oxidative capacity and insulin sensitivity of skeletal muscle .
these physiological benefits mainly occur according to the type and volume of the exercise , physical fitness of the mother , the time point in the pregnancy when the exercise is carried out and also the duration and intensity of the exercise [ 9 , 10 ] .
physical exercise training has been associated with reduced risk of metabolic disease and enhances both cardiorespiratory and metabolic functions [ 1114 ] .
these effects are mainly due to improved oxidative capacity and insulin sensitivity of skeletal muscle , since reduced muscle dysfunction and insulin resistance have been associated to the development of several chronic diseases [ 1520 ] .
exercise is considered moderate intensity when oxygen consumption is 5070% of the maximal oxygen consumption ( vo2max ) . in our previous study , we demonstrated in rats that a controlled moderate to low - intensity exercise before and during gestation ( 5 days week , 60  min day , 40%70% of vo2max ) increased the mothers ' resting oxygen consumption . in humans , the rate of placental bed blood flow increases , and more glucose and oxygen delivery to the placental site are observed in women submitted to a physical training .  
the effects of maternal moderate physical training can be expected modifying glucose metabolism in pregnancy once regular physical activity in nonpregnant individual is known to improve peripheral insulin sensitivity , glucose tolerance , and may aid in preserving -cell function .
regular practice of moderate exercise has been associated with improved fitness , reduction of body weight , and increased muscle mitochondrial biogenesis and antioxidant defence [ 5 , 23 ] .
our previous experimental data supports the idea that maternal physical training may protect mothers against perinatal undernutrition environment [ 21 , 24 ] .
recently , we showed in rats that physical training was able to normalise the effects of a low - protein diet on fasting serum glucose at the second and third weeks of gestation . this differential effect
observed in previously active compared with undernourished dams may be related to reduced risk of gestational diabetes mellitus , since an exercise program may improve insulin sensitivity and fasting plasma glucose concentrations of women at risk for gestational diabetes mellitus .  
thus , the main goal of this study was to evaluate the impact of a moderate to low intensity protocol of physical training during gestation on the secretion of insulin whose mothers were undernourished .
our hypothesis is that maternal physical training - induced physiological adaptations during gestation attenuate the impact of a perinatal low - protein diet on the secretion of insulin . this is a topic of particular interest as a maternal lifestyle
can be considered a therapeutic means of countering the effects of either maternal undernutrition or overnutrition .
experimental protocol was approved by the ethical committee of the biological sciences center , federal university of pernambuco , brazil and followed the guidelines for the care and use of laboratory animals .  
virgin female albino wistar rats ( rattus novegicus ) aged 60 days and weighting 180  11  g ( mean  s.e.m . ) were obtained from the department of physiology and biophysics , institute of biomedical sciences , university of so paulo , brazil .
female rats were maintained at a room temperature of 22  1c and controlled light - dark cycle ( dark 6:00 a.m.6:00 p.m. ) .
the standard laboratory chow for animals ( 52% carbohydrate , 21% protein , 4% lipids - nuvilab cr1-nuvital ) and water were given ad libitum .
animals were randomly divided into two groups : untrained rats ( nt , n = 10 ) and trained rats ( t , n = 10 ) .
trained rats were submitted to a training program of moderate running over a period of 4 weeks ( 5 days per week and 60 minutes per day ) on a treadmill ( millennium inbramed , brazil ) at a controlled intensity based on their vo2max. after a 4-week training period , the rats were mated ( 2 females for 1 male ) .
the day on which spermatozoa were present in a vaginal smear was designated as the day of conception ( day 0 of pregnancy ) .
half of rats of each group received either 17% casein diet or 8% casein ( low - protein group , lp ) isocaloric diet ad libitum ( table 1 ) .
thus , two more groups were formed : untrained ( c , n = 5 ) , trained ( t , n = 5 ) , untrained with low - protein diet ( lp , n = 5 ) , and trained with low - protein diet ( t + lp , n = 5 ) .
mother 's body weight was weekly determined throughout the experiment . on the time of delivery , the litter size and pup 's birth weight were recorded .
the litters of six pups represent the sample that was evaluated : control ( c , n = 5 ) ; trained ( t , n = 5 ) ; low - protein diet ( lp , n = 5 ) ; trained with low - protein diet ( t + lp , n = 5 ) .
the evaluation of body weight and gain of body weight of pups was recorded until the 3rd d of life with a marte scale with 100  mg precision .
considering that maternal physical exercise - induced changes depends on intensity , duration , and frequency of effort , our group standardized an experimental protocol of maternal physical training ( based on blood lactate concentration and oxygen consumption ) .
briefly , rats ran on a treadmill during the four weeks prior to pregnancy ( 5 days week and 60  min day , at 65% vo2max ) .
the protocol was divided into four progressive stages in each session : ( i ) warm - up ( 5 minutes ) ; ( ii ) intermediary ( 20 minutes ) ; ( iii ) training ( 30 minutes ) ; ( iv ) cool - down ( 5 minutes ) periods .
the percentage of vo2max during the sessions of training was kept around 5565% . during pregnancy ,
rats ran at a progressively lower intensity of effort ( 40% vo2max , 5 days week , and 20  min day ) until the 19th day of gestation .   mother 's body weight was daily recorded .
daily food consumption was determined by the difference between the amount of food provided at the onset of the dark cycle ( 06.00 hours ) and the amount of food remaining 24  h later .
body and food weights were recorded with a marte scale ( as-1000 ) with a 001-g accuracy .
islets were isolated from five fasting mothers in each group by collagenase digestion of pancreas followed by separation from pancreatic debris by centrifugation on ficoll gradients as previously described .
groups of five islets were initially incubated for 45  min at 37c in krebs - bicarbonate buffer containing 5.6  mm glucose and equilibrated with 95% o2/5% co2 , ph 7.4 .
the solution was then replaced with fresh krebs - bicarbonate buffer and the islets were incubated for a further 1  h with medium containing 5.6 or 16.7  mm glucose .
the incubation medium contained ( in mm ) : nacl 115 , kcl 5 , nahco3 24 , cacl2 2.56 , mgcl2 1 , and bsa 0.3% ( w / v ) .
the cumulative insulin release during 1  h was quantified by radioimmunoassay using rat insulin as standard .
data were analyzed by two - way repeated measures anova , with mothers ' diet and physical training as factors .
each litter of six pups was considered one sample , and statistical analyses were performed by using the mean values of each litter .
data analysis was performed using the statistical program graphpad prism 5 ( graphpad software inc . , la jolla , ca , usa ) .
body weight was lower in trained rats during the third and fourth weeks pregestation ( figure 1(a ) ) .
lp mothers showed a reduction in the body weight when compared to c mothers during gestation ( p < 0.01 ) , whereas physical training had no effect in the body weight ( figure 1(a ) ) .
the number of pups ( litter size ) born from mothers was similar among groups .
litter weight , birth weight , and body weight in the first three days of life of pups from mothers submitted to a low - protein diet during gestation , trained or not , were lower when compared to control ( table 2 ) .
trained mothers showed an increase of insulin secretion by isolated islets incubated with stimulatory ( 16.7  mm ) concentration of glucose ( figure 2 ) .
however , low - protein diet during gestation induced a reduction in the secretion of insulin by isolated islets when compared with both control and lp + t groups ( figure 2 ) .
maternal exercise guidelines preconize that 30  min , at least , of moderate - intensity exercise a day on most , if not all , days of the week is satisfactory for health . following these recommendations
, a controlled prospective studies have demonstrated that moderate pregestational exercise ( approximately 50% to 70% of vo2max ) is useful to increase metabolic rate ( reduction of body weight ) and improve cardiorespiratory fitness and maternal - fetal physiological reserve .
in addition , previous study has demonstrated an improved cardiovascular function , limited gain of body weight , and reduced risk of gestational diabetes mellitus and hypertension . in the present study , trained mothers demonstrated reduced body weight in the second and third weeks pregestation , but this normalized throughout gestation .
it is well established that regular physical exercise increases lean body mass and induces a higher utilisation of fatty acids as fuel by skeletal muscles once the intensity of the exercise is maintained at 6570% of vo2max .  
maternal physical training and low - protein diet did not alter the amount of food intake as seen in previous studies [ 25 , 33 ] .
however , dams submitted to a low - protein diet showed a reduction in body weight in the last week of gestation and the first days postdelivery .
maternal protein restriction is associated with lower stores of maternal nutrients , and it has been found previously that an undernutrition - induced reduction of maternal gain of body weight is positively correlated with a low birth weight , an impaired offspring growth rate , and a loss of lean mass during development .
duringpregnancy , there is an increase in basal hepatic glucose production , a progressive decrease in insulin sensitivity , and an associated 3.03.5-fold increase in insulin response in the last third of gestation .
herein we showed that maternal low - protein diet induced a reduction in the secretion of insulin .
our results are in accordance with previous studies where maternal low - protein ( 6% casein ) diet induced impairment in the metabolism of glucose and secretion of insulin [ 1 , 35 , 36 ] .
it has been suggested that the poor secretory response to glucose observed in islets from low - protein diet rats may be related to a defect in the ability of glucose to increase ca uptake and/or to reduce ca efflux from -cells   and a reduced islet mass and / orinsulinbiosynthesis .  
given that previous studies have described an inverse relationship between maternal insulin and maternal insulin sensitivity with fetal growth restriction , it is expected that the processes governing fetal growth will be also affected .
indeed , we and previous investigators have showed that pups from mothers submitted to a low - protein diet present a reduction in body weight at postnatal period [ 24 , 3739 ] .
these changes may result from a reduction in nutrient delivery to the fetus , then it would result in lower fetal insulin concentrations and a decrease in fetal igf - i and igf - ii that would downregulate fetus - placental growth .  
little is known about the effects of maternal physical activity on the secretion of insulin . here , we demonstrated for the first time that a controlled protocol of physical training before and during gestation increased pancreatic islets glucose - stimulatedinsulinsecretion .
this effect was particularly important once maternal protein - restriction has been shown to impair secretion of insulin leading to gestational diabetes as seen in previous study .
the underlying mechanism to the effects of maternal physical training on glucose - stimulated insulin secretion remains unknown but can be associated to increased pancreatic -cell mass by upregulation of growth and survival pathways ( akt , erk pathways ) in pancreatic islets .
in addition , it can be resulted from the high peripheral glucose uptake by skeletal muscle in response to moderate physical training .
indeed , our previous study demonstrated that dams submitted to a low - protein diet ( 8% casein ) during gestation present a higher fasting glycaemia than control ( 17% casein ) while trained mothers remained with normal glycaemia .
our data reinforce the impaired effects of a maternal low - protein diet on the pancreatic islets glucose - stimulatedinsulinsecretion that is preventable by moderate physical training .
the present study demonstrated that a controlled protocol of physical training during gestation resulted in increased insulin secretion , and mothers fed a low - protein diet during gestation showed similar results . this differential effect observed in
the present study refers to dams which were previously trained when were contrasted with pregnancy and low - protein diet .
these observations indicate that maternal physical exercise initiated in early pregnancy induces maternal adaptations and can be considered as a therapeutic mean of countering the effects of maternal undernutrition , which may provide a useful strategy since an exercise program may improve insulin sensitivity and fasting plasma glucose concentrations of women at risk for gestational diabetes .  
similarly to our results , previous study demonstrated that light - to - moderate physical activity during pregnancy is associated with reduced risk of abnormalglucosetolerance and hyperglycemia during pregnancy .
m / min , starting with a 20  min run and with a progressive daily increase of 5  min , throughout gestation showed a lower rise in blood glucose after an oral glucose load ( 2  g / kg body weight ) than untrainedpregnantrats . in human , previous study has found that associations between the physical activity and insulin sensitivity and -cell response do not appear to differ in pregnant versus nonpregnant women .
the present study showed that low - protein diet during gestation affects the secretion of insulin .
in contrast , pregestational and gestational moderate physical training acts as an environmental stimulus by increasing insulin secretion and attenuating the effects of perinatal undernutrition in rats , suggesting a potential strategy for compensation of insulin resistance and preventing gestational diabetes mellitus during late pregnancy .
 the effects of pregestational and gestational low - to - moderate physical training on insulin secretion in undernourished mothers were evaluated . 
 virgin female wistar rats were divided into four groups as follows : control ( c , n = 5 ) ; trained ( t , n = 5 ) ; low - protein diet ( lp , n = 5 ) ; trained with a low - protein diet ( t + lp , n = 5 ) . trained rats ran on a treadmill over a period of 4 weeks before mate ( 5 days week1 and 60  min day1 , at 65% of vo2max ) . at pregnancy , the intensity and duration of the exercise were reduced . 
 low - protein groups were provided with an 8% casein diet , and controls were provided with a 17% casein diet . at third day after delivery , mothers and pups were killed and islets were isolated by collagenase digestion of pancreas and incubated for a further 1  h with medium containing 5.6 or 16.7  mm glucose . 
 t mothers showed increased insulin secretion by isolated islets incubated with 16.7  mm glucose , whereas lp group showed reduced secretion of insulin by isolated islets when compared with both c and lp + t groups . 
 physical training before and during pregnancy attenuated the effects of a low - protein diet on the secretion of insulin , suggesting a potential role for compensation of insulin resistance and preventing gestational diabetes mellitus . 
1. Introduction
2. Methods
3. Results
4. Discussion
5. Conclusions