| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
Articles |
a Department of Biochemistry and Molecular Biology, Division of Reproductive Biology, and
b Departments of Psychology and Neuroscience, The Johns Hopkins University, Baltimore, Maryland 21205
 |
ABSTRACT |
|---|
 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
|
INTRODUCTION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Both spermatogenesis and steroidogenesis decline during testicular atrophy in seasonally-breeding mammals [1,3]. A variety of environmental factors such as photoperiod, food availability, temperature, and rainfall can be used as timing cues to initiate this reproductive quiescence, thus minimizing the costs of reproduction [4–7]. Photoperiod is the most salient cue for seasonally breeding species, and the effects of short (< 12 h light/day) days on testicular function have been well described in many seasonally breeding species [4,8].
Food restriction alone can inhibit both the maintenance and onset of reproductive capability; a nutritional threshold must be achieved, regardless of photoperiod, in order to prevent initiation of testicular regression and maintain full reproductive capabilities [9,10]. Interaction between photoperiod and food availability in the maintenance of male reproductive function also has been reported [9,11]. The extent of gonadal regression is often increased in the laboratory with exposure to multiple winter cues [10].
In white-footed mice (P. leucopus), testicular regression induced by short day lengths is a result of apoptosis. After 4–10 wk of exposure to a short (LD 8:16) photoperiod, the extent of testicular apoptotic DNA fragmentation increases, as measured by in situ terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) and DNA laddering [12]. Apoptosis involves a regulated cascade of morphological and biochemical changes leading to DNA fragmentation and, ultimately, the efficient removal of the targeted cell [13–16]. Apoptosis can be recognized morphologically by characteristic membrane blebbing, condensation of the nucleus, DNA fragmentation, and formation of the apoptotic bodies that are cleanly phagocytosed by adjacent cells [15,16]. In contrast, necrotic, or pathophysiological cell death can be identified by large swollen nuclei, irregular loss of membrane integrity, and evidence of inflammation due to cell lysis [17]. Localized necrotic cell death in the testis causes collapsed seminiferous tubules and excessive germ cell desquamation that often leads to vacuolization in the seminiferous epithelium [17,18].
Dietary restriction provides both an acute cellular stressor as well as a cue for seasonal testicular regression [4,9]. As yet, it is not known whether mechanisms that mediate testicular regression as a result of restricted caloric intake are similar to the apoptotic-regulated regression induced by short day lengths, or the result of unorganized necrotic cell death [12]. We show here that reduced food intake in fact induces testicular regression that is mediated by apoptosis and, further, that there is a significant interactive effect of photoperiod and food restriction on apoptosis.
|
MATERIALS AND METHODS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Eighty adult (> 60 days of age) male white-footed mice (P. leucopus) were obtained from the Peromyscus Stock Breeding Center at the University of South Carolina (Columbia, SC). Animals were housed individually in polypropylene cages (28 x 7.5 x 13 cm) at 21 ± 2°C and 50 ± 5% relative humidity. All experiments were conducted in our AAALAC-approved facilities, in accordance with NRC guidelines for use of laboratory animals. Tap water was available ad libitum (ad lib) for the duration of the experiment. Ad lib mice used in this study include randomly-selected (formalin-fixed) animals from a complementary study in our laboratory [12]. All mice were housed in photoperiodic conditions during overlapping periods of time. Prior to the experiment, mice were individually housed and allowed to acclimate for 5 days, and baseline food consumption was assessed daily for 7 days by weighing blocks of food (Agway Prolab 2000, Syracuse, NY) in each hopper. Mice were housed in long (16L:8D) or short (8L:16D) photoperiods and fed either ad lib or 70% of the average baseline food intake for 2, 4, 6, or 8 wk (n = 5 per group).
Experimental Protocol
Testosterone RIA At the end of each photoperiod exposure, terminal blood samples were collected into iced heparinized tubes from animals under light methoxyflurane anesthesia (Metofane; Schering-Plough, Union, NJ), and centrifuged for 30 min at 2500 rpm at 4°C. After separation, plasma was stored at -80°C until plasma hormone values were determined in duplicate by a single RIA using 125I kits (ICN Biomedicals, Costa Mesa, CA). The ICN testosterone assay is highly specific; cross reactions with other steroids are < 0.1%–0.78%. The 125I kit has been validated for use in small rodents in our laboratory [19]. The intraassay variation is 4.43%, and interassay variation is 6.42%.
Tissue processing Following collection of the terminal blood sample, the left testis was freed from the epididymis, weighed, snap-frozen in a bath of dry ice/ethanol, and stored at -80°C. Animals were then perfused through the heart with 50 ml 0.9% saline followed by 200 ml of 10% neutral buffered formalin (Electron Microscopy Sciences, Fort Washington, PA) as a fixative. After fixation, the right testis was removed, weighed, and postfixed in 10% formalin for 5 days. Tissue was then washed in PBS and dehydrated in 70% ethanol prior to paraffin embedding.
For TUNEL labeling, 6-µm sections were collected every 50 µm of tissue and stained for apoptotic activity (TACS 2TdT; Trevigen, Gaithersburg, MD). Twelve cross sections per testis were stained and completely counted. Cells that incorporated the labeled biotinylated nucleotides were considered to be TUNEL-positive (apoptotic) and were counted under bright-field illumination (40x) on a Zeiss Axioplan 2 microscope (Thornwood, NY) using Stereoinvestigator software (Microbrightfield, Colchester, VT). Negative control sections, processed without TdT, showed no staining. Control slides (Trevigen) were processed with experimental slides and served as a positive control for apoptotic signal. Apoptotic activity was quantified by counting the number of cells positive for TUNEL staining within each testis cross section. The section thickness and intersection intervals that were used prevented the double-counting of labeled cells. To control for reduction of testis size, the number of TUNEL-positive cells was expressed as number of apoptotic cells in each testis cross section per total number of seminiferous tubules within that cross section, as we have done previously [12].
To determine reproductive competence, spermatogenic activity was evaluated in 10 seminiferous tubules per animal using the spermatogenic index developed by Grocock and Clarke [20]. The spermatogenic index is a comparative measure of the extent of spermatogenesis in the seminiferous epithelium. Scores assigned ranged from 1–5. A value of ``5" was given to large tubules displaying complete spermatogenesis; a score of ``1" was assigned to small tubules that contained primarily Sertoli cells, spermatogonia, and primary spermatocytes.
DNA isolation and analysis
To determine the extent of TUNEL labeling, DNA from food-restricted males was isolated from 0.05–0.10 g of fresh-frozen pulverized testicular tissue. The tissue was lysed in a buffer containing 0.2% Triton X-100, 10 mM Tris-HCL, and 10 mM EDTA and then lightly homogenized and extracted with phenol/chloroform/isoamyl alcohol. After precipitation with sodium acetate and isopropanol, total DNA was quantified spectophotometrically based on absorbance at 260 nm. Aliquots of DNA (0.5 µg) were processed for 3' end labeling with [
-32P]dideoxy-ATP (10 µCi/µl; ICN Biomedicals) using 5 U/µl Klenow enzyme (Trevigen). Labeled samples were separated on a 1.5% Trevigel, dried without heat for 2 h on a gel dryer (Bio-Rad, Hercules, CA), and exposed to Hyperfilm-MP (Amersham Life Sciences, Arlington Heights, IL) at -80°C for 18 h. After autoradiography, lanes were excised from the gel, and areas that corresponded to low (< 15 kb) and high (> 15 kb) molecular weight DNA fractions were counted in a ß-counter to determine the extent of apoptotic activity.
Statistical Analysis
Statistical evaluation of mean differences between experimental groups was performed by ANOVA and, for measurements lacking equal variance, with a Kruskal-Wallis ANOVA on Ranks using the Sigma Stat software package (Jandel Scientific, San Rafael, CA). To isolate significant differences between groups, the Student-Newman-Keuls method was used for the pairwise multiple comparisons. Mean differences were considered statistically significant when P < 0.05.
|
RESULTS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
The body mass of white-footed mice was not significantly affected by photoperiod or mild food restriction under the conditions of this study. The average body mass for long-day ad lib-fed males was 20.5 ± 0.5 g; short-day ad lib-fed males, 20.2 ± 0.5 g; long-day food-restricted animals, 18.7 ± 0.5 g; and short-day food-restricted males was 19.9 ± 0.6 g.
Testis Mass
Figure 1A shows the effect of photoperiod on paired testis mass among mice fed ad lib. No changes in relative (mg paired testis mass/g body mass) (data not shown) or absolute paired testis mass (Fig. 1A) were observed through 8 wk of long (16L:8D) day exposure. In all cases, absolute and relative testis mass showed similar patterns of significant mean difference; because the pattern of results are similar and for the sake of brevity, only absolute paired testis mass results are reported here. Among ad lib-fed mice, paired testis mass was significantly reduced (38%) by 8 wk of short (8L:16D) photoperiods.
|
Restriction of food intake affected testis mass in mice housed either in long (Fig. 1B) or short (Fig. 1C) photoperiods. Among mice housed in long days (Fig. 1B), a decrease in paired testis mass was observed after 8 wk of food restriction. At that time, testis mass was reduced by 27% in food-restricted as compared to ad lib-fed mice. The interaction of food restriction and short-day exposure produced the maximal reduction in paired testis mass (Fig. 1C). Short-day exposure in ad lib-fed mice induced a decrease in testis mass at 8 wk, whereas significant gonadal regression was detected at 6 wk in short-day food-restricted mice (Fig. 1C). At 8 wk, paired testis mass of food-restricted mice housed in short days was reduced 22% compared with ad lib-fed short day mice, 34% as compared with food-restricted mice housed in long days, and 52% when compared with ad lib-fed mice housed in long days.
Testosterone
Plasma testosterone concentrations did not change among long-day (16L:8D) white-footed mice fed ad lib (Fig. 2A). Among ad lib-fed mice exposed to short (8L:16D) photoperiods, testosterone concentrations were reduced at 6 and 8 wk. At 8 wk, ad lib-fed males had a 48% decrease in plasma testosterone concentrations in short as compared with long-photoperiod exposure (Fig. 2A, post hoc analysis: P = 0.017). In contrast to plasma testosterone concentrations among long-day males fed ad lib, long-day mice that had been food-restricted exhibited a 42% decrease of plasma testosterone at 8 wk compared with 8-week, long-day, ad-lib fed mice (Fig. 2B).
|
Food restriction interacted with short photoperiod to maximally decrease plasma testosterone concentrations (Fig. 2C). Thus, whereas short photoperiod exposure alone resulted in a significant decrease in plasma testosterone concentrations at 6 wk, the combined exposure to short photoperiod and food restriction induced a decrease in testosterone concentrations at 4 wk as compared with ad-lib short-day mice at the same time point (Fig. 2C). At 4 wk of short-day exposure, a significant decrease in plasma testosterone was recorded in food-restricted males as compared with short-day males fed ad lib for 2 and 4 wk, and short-day males food-restricted for 2 wk. At 6 or 8 wk of short-day exposure and food restriction, plasma testosterone concentrations were reduced as compared with mice in all ad lib-fed groups and 2-week food-restricted males (Fig. 2).
Spermatogenic Index
Spermatogenic index (SI), a measure of reproductive capacity [20], did not differ among long-day mice fed ad lib; the average spermatogenic index value was 4.8 ± 0.1 in ad lib-fed long-day males and 4.4 ± 0.1 in short-day males. The effect of short-day as compared with long-day photoperiod was significant only at 8 wk of short photoperiod exposure (SI = 3.9 ± 0.2). The mean SI value for food-restricted long-day mice (4.2 ± 0.1) did not differ from the average long-day ad-lib SI until week 8 of restricted food intake (SI = 3.4 ± 0.1). Spermatogenic index values decreased with simultaneous exposure to short photoperiods and restricted food intake; the average SI for short-day, food-restricted males was 3.4 ± 0.4, maximal differences in comparison with short-day, ad lib-fed males occurred at 6 and 8 wk of short photoperiod and food restriction exposure (SI = 2.2 ± 0.6, and 2.9 ± 0.6, respectively).
Apoptotic DNA fragmentation
In situ end labeling TdT-mediated end-labeling of fragmented DNA (TUNEL) was used on testis cross sections to assess apoptosis in individual cells. Examples of results obtained are shown in Figure 3. Comparatively few TUNEL labeled-cells were seen in long-day males fed ad lib (Fig. 3A,B). In contrast, there was an obvious increase in apoptotic labeling after exposure to the combination of short photoperiod and food-restriction (Fig. 3C,D). Based on their observed position within the seminiferous epithelium and the size and appearance of the nuclei, TUNEL-labeled cells were identified predominantly as spermatocytes. Spermatids and spermatogonia rarely labeled. Examination of peritubular connective tissue and endothelial cells revealed little to no apoptotic staining. In testes of both short and long-day males, features characteristic of necrotic cell death such as swollen germ cells and vacuolization were rarely noted. These observations were too infrequent to allow statistical comparisons.
|
The number of apoptotic cells within each testis cross section was quantified (number of TUNEL-positive cells per seminiferous tubule) (Fig. 4). No differences in the number of stained cells were observed among ad lib-fed males housed in long days through 8 wk (Fig. 4A); exposure to short photoperiod resulted in a significant increase in the number of apoptotic cells in ad-lib fed mice at 4, 6, and 8 wk. The increase in TUNEL labeling at 4, 6, and 8 wk of short-day exposure in ad lib males was significant in comparison to long-day ad lib mice at all time points assayed.
|
Food restriction during long-day exposure caused an initial increase in apoptotic labeling at 2 and 4 wk compared with ad lib-fed mice (Fig. 4B); at 6 and 8 wk, however, the numbers of apoptotic cells were equivalent in ad lib and restricted mice. Short-day food-restricted males showed increases in TUNEL labeling as early as 2 wk, and at 4, 6, and 8 wk in comparison with short-day ad lib-fed males (Fig. 4C). Restricting food intake for 6 and 8 wk in short-photoperiod resulted in the maximal number of apoptotic cells when compared with all other experimental groups.
DNA fragmentation Figure 5 shows agarose gel electrophoresis of DNA extracted from the testes of food-restricted males housed for 2–8 wk in long (16L:8D) and short (8L:16D) photoperiods. No apparent increases in low molecular weight (< 15 kb) apoptotic DNA fragments were noted in long-day food-restricted males at 2, 4, or 6 wk, but there was an apparent small increase in laddering noticeable at 8 wk. In contrast, an increase in the degree of low molecular weight DNA was observed as early as 2 wk in the short-day food-restricted males, and there were increases thereafter through 8 wk. These results are similar to those obtained by TUNEL staining (Fig. 5A).
|
The effect of photoperiod on DNA fragmentation in food-restricted mice was quantified with ß-scintillation counting of the [-32P]dideoxy-ATP-labeled testicular DNA (Fig. 5B). No significant change in percent of low molecular weight DNA was detected among long-day food-restricted mice (Fig. 5B). An increase in percent of low molecular weight DNA per total lane DNA was observed after exposure to 4, 6, and 8 wk of short-day exposure in food-restricted mice when compared with food-restricted males housed in long days for 2, 4, and 6 wk (Fig. 5B, P < 0.05). The maximal degree of total testis DNA laddering was observed at 8 wk of short-day exposure and food restriction; the apoptotic fragmentation in 8 week short-day males was increased in comparison with long-day food-restricted males at every time point assayed, as well as with males housed in short days for 2 wk.
Overall, there was a significant negative correlation between TUNEL-positive cells and paired testis mass in both ad lib-fed and food-restricted males exposed to short and long photoperiods (r2 = -0.14, P = 0.002; and r2 = -0.16, P = 0.012, respectively). Plasma testosterone concentrations in both ad lib-fed and food-restricted males in short and long days also correlated negatively with the number of TUNEL-labeled cells (r2 = -0.16, P < 0.01; and r2 = -0.20, P = 0.05, respectively). The spermatogenic index, a measure of reproductive competence, also correlated negatively with the number of cells positively labeled for apoptosis in ad lib and food-restricted mice in short and long photoperiods (r2 = -0.35, P < 0.001; r2 = -0.71, P < 0.01, respectively).
|
DISCUSSION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
All other things being equal, restricted food intake would be expected to result in reduced body weight [5,22–23]. The 30% food restriction in the present study, however, reflects a relatively mild alteration in daily food intake. Previous studies from our lab have indicated that mild restriction does not affect body mass at 8 wk; however, reproductive function is reduced after 8 wk in 16L:8D and 4 wk in 8L:16D [7,11]. Other studies, however, have reported body mass reductions after similar levels of food restriction [22]. The explanation for this discrepancy remains uncertain; however, it has been suggested that a decrease in food intake may result in a decline in locomotor activity in Peromyscus species [5]. Reduced activity levels could potentially prevent alterations in body mass in mildly food-restricted mice.
Short-day induced reproductive inhibition is accelerated by restricted food intake [8,11,23–24]. Compared with all other experimental groups in the present study, short-day food-restricted mice exhibited maximal decreases in testis mass and plasma testosterone concentrations, coupled with the largest increases in apoptotic cell death. This increase in apoptosis was first observed at 2 wk of food restriction and short-day exposure and reached nearly 5-fold by 8 wk. In contrast, increases in apoptosis were seen later (at 4 wk) and less extensively in ad lib-fed short-day mice. The increase in testicular apoptosis after simultaneous limitation of food intake and short-day exposure agrees with the accelerated reproductive inhibition observed in deer mice (P. maniculatus) and golden hamsters (Mesocricetus auratus) after exposure to these combined cues [11,23–24]. These results indicate that males in environments that more closely simulate natural winter conditions may depend on multiple factors to undergo complete testicular regression.
Peak apoptotic activity in both food-restricted and ad lib-fed short-day mice occurred at 6 and 8 wk of experimental exposure, at time points when plasma testosterone concentrations had significantly decreased. Interestingly, the initial reductions in plasma testosterone concentrations occurred after the first detected increases in testicular apoptosis. This is consistent with the results of previous studies in which reduction in testosterone occurred after initial increases in testicular apoptosis in ad lib-fed adult white-footed mice and peri-pubertal hamsters [12,21]. This suggests that low testosterone concentrations may not mediate the onset of testicular regression induced by food restriction and short photoperiods, though a reduction in plasma testosterone concentration may accelerate the regression process. Alternatively, it is also possible that intra-testicular testosterone concentrations decrease before serum testosterone concentrations decline, and before increased apoptotic cell death. This remains a possibility because a careful analysis of intra-testicular as compared with plasma testosterone concentrations during testicular regression has not yet been conducted in this species.
The increase in testicular apoptotic cell death after exposure to short photoperiod or short photoperiod plus food restriction, could potentially result from decreased gonadotropin secretion. Withdrawal of gonadotropins induces increases in apoptotic germ cell death [25–28]. Decreasing concentrations of FSH in response to either specific immunoneutralization of FSH or transfer to short photoperiod are concomitant with increases in apoptotic cell death in adult and peripubertal hamsters, respectively [21,27]. Plasma concentrations of LH also decrease in both short-photoperiod and food-restricted mice [4]. Immunization against LH induces an increase in apoptotic germ cell death in rats, and exogenous administration of hCG to hypophysectomized rats reduces the amount of DNA fragmentation that occurs with gonadotropin withdrawal [26–28]. Reduction in gonadotropin signaling to the testis during testicular regression may induce Sertoli cell-mediated germ cell death, potentially through the Fas death receptor system [12].
The energetically-conservative process of accelerated gonadal regression in response to food restriction, short photoperiods, and both factors simultaneously, as observed in the present study, appears to be mediated by an increased presence of apoptotic cell death within the testis. Winter-typical conditions, such as limited access to high-quality food, decrease available metabolic energy in small rodents [29]. Food restriction can be an extreme metabolic stressor; restricted food intake shifts resources away from reproductive function and towards somatic cell maintenance [30–32]. Gonadal regression may counteract this decrease in available energy, generating metabolic benefits in deer mice (P. maniculatus) [32]. Although the cascade of events that characterizes apoptosis has been postulated to require substantial cellular energy, death via necrotic mechanisms induces cellular lysis, a process that necessitates an energetically-expensive immune response [33–34]. Additional studies of testicular atrophy in response to severe caloric restriction are necessary to determine fully the roles of apoptotic and necrotic cell death in the testis during metabolic stress.
|
ACKNOWLEDGMENTS |
|---|
|
FOOTNOTES |
|---|
1 This work was supported by NIMH grant 57535 (R.J.N.), NICHHD Training Grant T32-HD-07276 (K.A.Y.), and Population Center Grant P30-HD 06268 (B.R.Z). 
2 Correspondence: Kelly A. Young, Department of Biochemistry and Molecular Biology, Division of Reproductive Biology, Johns Hopkins University School of Hygiene and Public Health, 615 N. Wolfe St., Baltimore, MD 21205. FAX: 410 516 6205; kyoung{at}jhsph.edu
Accepted: September 28, 1999.
Received: May 13, 1999.
|
REFERENCES |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
This article has been cited by other articles:
|
|
 |
|
  M Carvalho, L Mateus, F Afonso, S Van Harten, L Alfaro Cardoso, D A Redmer, and G Ferreira-Dias Testicular angiogenic activity in response to food restriction in rabbits Reproduction, March 1, 2009; 137(3): 509 - 515. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. S. Rocha, M. S. Bonkowski, L. R. Franca, and A. Bartke Mild Calorie Restriction Does Not Affect Testosterone Levels and Testicular Gene Expression in Mutant Mice Experimental Biology and Medicine, September 1, 2007; 232(8): 1050 - 1063. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. V. Teixeira, D. Silandre, A. M. de Souza Santos, C. Delalande, F. J B Sampaio, S. Carreau, and C. da Fonte Ramos Effects of maternal undernutrition during lactation on aromatase, estrogen, and androgen receptors expression in rat testis at weaning J. Endocrinol., February 1, 2007; 192(2): 301 - 311. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. Morales, C. Ferrer, A. Zuasti, J. C. Garcia-Borron, M. Canteras, and L. M. Pastor Apoptosis and Molecular Pathways in the Seminiferous Epithelium of Aged and Photoinhibited Syrian Hamsters (Mesocricetus auratus) J Androl, January 1, 2007; 28(1): 123 - 135. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. M Pyter, A. K Hotchkiss, and R. J Nelson Photoperiod-induced differential expression of angiogenesis genes in testes of adult Peromyscus leucopus Reproduction, February 1, 2005; 129(2): 201 - 209. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. Fu, L. Xi, Y. Wu, R. McCarter, A. Richardson, M. Hickey, and E.-S. Han Hepatic Genes Altered in Expression by Food Restriction Are Not Influenced by the Low Plasma Glucose Level in Young Male GLUT4 Transgenic Mice J. Nutr., November 1, 2004; 134(11): 2965 - 2974. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. A. Young, B. R. Zirkin, and R. J. Nelson Testicular Apoptosis Is Down-Regulated during Spontaneous Recrudescence in White-Footed Mice (Peromyscus leucopus) J Biol Rhythms, October 1, 2001; 16(5): 479 - 488. [Abstract] [PDF]
|
|
|
|
 |
|
 K. A. Young, G. F. Ball, and R. J. Nelson Photoperiod-Induced Testicular Apoptosis in European Starlings (Sturnus vulgaris) Biol Reprod, February 1, 2001; 64(2): 706 - 713. [Abstract] [Full Text]
|
|
|
|
 |
|
 K. A. Young and R. J. Nelson Short photoperiods reduce vascular endothelial growth factor in the testes of Peromyscus leucopus Am J Physiol Regulatory Integrative Comp Physiol, September 1, 2000; 279(3): R1132 - R1137. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
Significant difference as compared with ad lib-fed mice at 4 and 6 wk of short days. *Significant difference as compared with short-day mice fed ad lib for 2, 4, and 6 wk and to week-2 food-restricted mice housed in short days
