J. Anim. Sci. 2005. 83:586-596
© 2005 American Society of Animal Science
ANIMAL GROWTH, PHYSIOLOGY, AND REPRODUCTION |
Influence of nutrient intake and body fat on concentrations of insulin-like growth factor-I, insulin, thyroxine, and leptin in plasma of gestating beef cows1
C. A. Lents*,2,
R. P. Wettemann*,3,
F. J. White*,4,
I. Rubio*,
N. H. Ciccioli*,
L. J. Spicer*,
D. H. Keisler
and
M. E. Payton
* Department of Animal Science, Oklahoma Agricultural Experiment Station, Stillwater 74078;
and
Department of Animal Science, University of Missouri, Columbia 65211; and
and
Department of Statistics, Oklahoma State University, Stillwater 74078
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Abstract
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Pregnant Angus x Hereford cows (n = 73) were used to determine the effects of amount of nutrient intake and BCS on concentrations of IGF-I, insulin, leptin, and thyroxine in plasma. At 2 to 4 mo of gestation, cows were blocked by BCS and assigned to one of four nutritional treatments: high (H = a 50% concentrate diet fed ad libitum in a drylot) or adequate native grass pastures and one of three amounts of a 40% CP supplement each day (M = moderate, 1.6 kg; L = low, 1.1 kg; or VL = very low, 0.5 kg; as-fed basis). After 110 d of treatment, all cows grazed dormant native grass pasture and received 1.6 kg/d of a 40% CP supplement. At 68, 109, and 123 d of treatment, cows were gathered, and plasma samples were collected by tail venipuncture (fed sample). After 18 h without feed and water, a second plasma sample was collected (fasted sample). At 109 d of treatment, BCS was greatest (P < 0.05) for H cows, similar for M and L cows, and least for VL cows. Concentrations of insulin and leptin were greater (P < 0.05) for H cows than for M and VL cows at 68 and 109 d, but similar for all groups at 123 d. Thyroxine in plasma was greatest (P < 0.05) for H cows at 68 d and similar for cows on all treatments at 123 d. Concentrations of IGF-I, insulin, and leptin in fed and fasted cows were positively correlated with BCS at 109 d. Body condition was predictive of concentrations of IGF-I, insulin, and leptin when cows had different nutrient intakes, but BCS accounted for less than 12% of the variation in plasma concentrations of IGF-I, insulin, and leptin when nutrient intake was the same for all cows. We conclude that amount of nutrient intake has a greater influence than body energy reserves on IGF-I, insulin, and leptin concentrations in the plasma of gestating beef cows.
Key Words: Beef Cows • Body Condition • Insulin • Insulin-Like Growth Factor I • Leptin • Nutrition
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Introduction
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Nutrient intake and body energy reserves regulate reproductive function of cattle (Wettemann et al., 2002
; Diskin et al., 2003). Cows with greater body condition at parturition resume estrous cycles sooner than thin cows (Richards et al., 1986; Houghton et al., 1990; Spitzer et al., 1995). If nutrient intake is limited and body condition is inadequate, cows (Richards et al., 1989a) and heifers (Rhodes et al., 1996; Bossis et al., 1999) cease to ovulate; however, the signal(s) by which the hypothalamus, pituitary, and ovary sense changes in nutritional status or body energy reserves is not fully established.
Decreased nutrient intake by cows results in inadequate fat reserves and decreased serum concentrations of IGF-I (Houseknecht et al., 1988; Granger et al., 1989; Richards et al., 1991), insulin (Trenkle, 1978; Vizcarra et al., 1998; Bossis et al., 1999), and thyroxine (Richards et al., 1995). Decreased concentrations of IGF-I in plasma are associated with increased postpartum anestrous intervals in cows (Nugent et al., 1993; Ciccioli et al., 2003).
Leptin is synthesized and secreted by adipose tissue, and leptin concentrations in plasma are positively correlated with the amount of body fat in sheep (Delavaud et al., 2000) and cattle (Delavaud et al., 2002). Nutrient intake causes dramatic alterations in plasma leptin in cattle (Amstalden et al., 2000; Ciccioli et al., 2003), and leptin stimulates LH secretion in undernourished beef cows (Amstalden et al., 2002; Zieba et al., 2003). It has not been established whether the amount of adiposity is the primary regulator of concentrations of leptin in cattle or whether nutrient intake influences concentrations of leptin in plasma independent of the amount of body fat. The objective of the present study was to determine the effects of body fat reserves and amount of nutrient intake on plasma concentrations of IGF-I, insulin, leptin, and thyroxine in mature pregnant beef cows.
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Materials and Methods
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Animals and Experimental Procedures
The Institutional Animal Care and Use Committee of Oklahoma State University approved all animal-related procedures used in this study. Mature pregnant Angus x Hereford cows (n = 73; BW = 514 ± 15 kg) were used to determine the effects of body condition and nutrient intake on concentrations of IGF-I, insulin, leptin, and thyroxine in plasma. At 112 ± 15 d of gestation (September 28), cows were blocked by BCS (1 = emaciate, and 9 = obese; Wagner et al., 1988) and assigned to one of four nutritional treatments. Cows assigned to the high (H) treatment (n = 13) were maintained in a drylot, with ad libitum access to a high-energy diet (11.1% CP, and 1.16 Mcal of NEm and 0.9 Mcal of NEg/kg of DM). The diet comprised (DM basis) rolled corn (39.7%), ground alfalfa pellets (35.5%), cottonseed hulls (22%), cane molasses (2.5%), and salt (0.3%). The remaining cows grazed native grass pasture with adequate forage and received one of three amounts of a 40% CP supplement (as-fed basis) each day: moderate (M; 1.6 kg of supplement, n = 12), low (L; 1.1 kg of supplement, n = 24), or very low (VL; 0.5 kg of supplement, n = 24). Nutritional treatments were used to produce cows with a range of BCS. Cows on different treatments were grouped in similar pastures, and cows were rotated among pastures every 2 wk. After 110 d of treatment, all cows grazed the same dormant native grass pasture with adequate forage and received 1.6 kg/d of a 40% CP supplement (as-fed basis). Cows were gathered on 68, 109, and 123 d of treatment at 1500, and within 1 h, a plasma sample was collected from each cow by tail venipuncture (fed sample). Cows were restricted from feed and water for 18 h, and a plasma sample was collected to evaluate the effect of fasting (fasted sample). Body condition score was determined at the start of treatment and at 68, 109, and 123 d.
Hormone Analyses
Concentrations of IGF-I in plasma were quantified by RIA after acid ethanol extraction (16 h at 4°C; Echternkamp et al., 1990), and recombinant human IGF-I (R& D Systems, Minneapolis, MN) was used as the standard. Intra- and interassay CV were 10 and 14%, respectively. Concentrations of insulin in plasma were quantified by a solid-phase RIA for human insulin (Bossis et al., 1999; Diagnostic Products Corp., Los Angeles, CA) with bovine pancreatic insulin as the standard (Sigma Chemical Co., St. Louis, MO). Intra- and interassay CV were 11 and 16%, respectively. Concentrations of leptin in plasma were determined by an RIA specific for ovine leptin and validated for use with bovine plasma (Delavaud et al., 2000). All samples were quantified in one assay, and the intraassay CV was 8%). Concentrations of thyroxine in plasma were quantified with a solid-phase RIA for human thyroxine (Diagnostic Products Corp.) validated for use with bovine plasma (Ciccioli et al., 2003). All samples were quantified in one assay, and the intraassay CV was 13%. Concentrations of IGF-I, insulin, and leptin were quantified in samples collected on 68, 109, and 123 d. Concentrations of thyroxine were determined in samples collected on 68 and 123 d.
Statistical Analyses
Mixed-model ANOVA procedures of SAS (SAS Inst., Inc., Cary, NC) for repeated measures were performed for each specified day of treatment to determine effects of nutritional treatment (H, M, L, and VL) on concentrations of hormones. Access to feed (fed or fasted) was used as the split-unit factor. Diets were used to create cows with different BCS, and diet and pasture were confounded. Data were collected for individual cows to evaluate the effect of BCS on concentrations of hormones in plasma. Because cow was the experimental unit for treatment, cow nested within treatment was used as the error term to test treatment effects, whereas the pooled residual was used as the error term to test access to feed and its interaction with treatment. Because a split-plot analysis was used, the within-cow covariance structure was, by default, compound symmetric. Because analysis for repeated measures was used, degrees of freedom for the pooled error term were calculated using Satterthwaite’s approximation. Orthogonal contrasts were used to compare treatment means. If there was a significant (P < 0.05) interaction between treatment and access to feed, orthogonal contrasts were used to compare treatment means within fed and within fasted samples. Additionally, if the interaction of treatment and access to feed was significant, means between fed and fasted samples, within a treatment, were compared using the SLICE option of the LSMEANS statement of SAS.
Simple correlation analyses were used to determine linear relationships of BCS and concentrations of IGF-I, insulin, leptin, and thyroxine in plasma after cows were fed or fasted. Regression analyses using first-, second-, or third-order models were used to evaluate any potential curvilinear relationship of BCS and concentrations of IGF-I, insulin, leptin, or thyroxine in plasma of fed and fasted cows on a specified day. Second- or third-order factors for BCS were included in the model if they had a P-value of <0.10 and the R2 of the model was increased by 
2%. The relationships of BCS and hormones in plasma were fit using separate models for fed and fasted samples. Indicator regression analysis was used to determine whether slopes of fed and fasted regression lines were different (Neter et al., 1989; Steel et al., 1997).
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Results
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Effects of Nutrient Intake
At the initiation of treatments, BCS did not differ (P = 0.90) for any of the treatment groups (4.6 ± 0.1) due to blocking. After 68 d of treatment, BCS was greatest (P < 0.01) for H cows (6.9 ± 0.09) and similar for M, L, and VL cows (5.0 ± 0.1, 5.1 ± 0.1, and 4.9 ± 0.1, respectively). Differences in BCS among cows on different treatments were the result of diet, pasture, or the interaction of diet x pasture, and diet and pasture were confounded. The effects of BCS on plasma hormones were evaluated in this experiment. There was no evidence that cows respond differently depending on the diet used to achieve a BCS. There was a treatment x access to feed interaction for concentrations of IGF-I (P < 0.001) and insulin (P < 0.01) at 68 d of treatment. In plasma obtained after cows had access to feed and water, or were fasted, concentrations of insulin were greatest (P < 0.001) for H cows compared with cows on all other treatments (Figure 1B), but concentrations of IGF-I were not influenced by treatment (Figure 1A). Low cows had greater (P < 0.01) concentrations of insulin in fasted samples than M or VL cows (Figure 1B). Fasting of L and VL cows (Figure 1A) did not influence concentrations of IGF-I; however, IGF-I concentrations in fasted samples were greater (P < 0.01) for H cows and less (P < 0.05) for M cows compared with fed samples (Figure 1A). Concentrations of insulin in samples from H and M cows after fasting were less (P < 0.05) than in samples when cows had access to feed and water (Figure 1B). Fasting in L and VL cows did not influence insulin concentrations. Concentrations of leptin in plasma after 68 d were not influenced by fasting or treatment x fasting, but there was a treatment effect (P < 0.01). High cows had greater concentrations of leptin in fed (P < 0.05) and fasted (P < 0.01) samples compared with all other treatments (Figure 1C). Concentrations of thyroxine in plasma were not affected by access to feed, but H cows had greater (P < 0.05) thyroxine than cows on all other treatments (Figure 2A).
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Figure 1. Least squares mean concentrations of IGF-I (A), insulin (B), and leptin (C) in plasma of cows at 68 d of nutritional treatments: high (H = a 50% concentrate diet fed ad libitum in a drylot), or adequate native grass pastures and one of three amounts of a 40% CP supplement each day (M = moderate, 1.6 kg; L = low, 1.1 kg; or VL = very low, 0.5 kg; as-fed basis). a,b,cWithin fed or fasted cows, bars without a common letter differ, P < 0.05. y,zWithin a treatment, bars without a common letter differ, P < 0.05. Standard errors were 3.8, 0.06, and 0.4 for IGF-I, insulin, and leptin, respectively.
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Figure 2. Least squares mean concentrations of thyroxine in plasma of cows at 68 d (A) and 123 d (B) of nutritional treatments: high (H = a 50% concentrate diet fed ad libitum in a drylot), or adequate native grass pastures and one of three amounts of a 40% CP supplement each day (M = moderate, 1.6 kg; L = low, 1.1 kg; or VL = very low, 0.5 kg; as-fed basis). a,bWithin fed or fasted cows, bars without a common letter differ, P < 0.05. Standard errors were 1.7 and 1.8 for thyroxine at 68 and 123 d of treatment, respectively.
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At 109 d of treatment, BCS was greatest (P < 0.01) for H cows (6.7 ± 0.1). Body condition score was similar for M and L cows (4.8 ± 0.1 and 5.0 ± 0.1, respectively), and BCS was less (P < 0.05) for VL cows (4.7 ± 0.1) than for L cows. There were interactions between treatment and access to feed on concentrations of IGF-I (P < 0.01), insulin (P < 0.01), and leptin (P < 0.001) in plasma at 109 d of treatment. High cows had greater concentrations of IGF-I in both fed (P < 0.05) and fasted (P < 0.001) samples than M or VL cows but did not differ from L cows (Figure 3A). Concentrations of IGF-I were greater in fed samples than fasted samples for H cows (P < 0.001), M cows (P < 0.001), and L cows (P < 0.05), but were similar for VL cows at 109 d (Figure 3A). Concentrations of IGF-I were greater for M and L cows in fed (P < 0.05) and fasted (P < 0.001) samples than for VL cows (Figure 3A). Concentrations of insulin in samples from fed cows were greater for H and L cows compared with all other cows (P < 0.001; Figure 3B). Concentrations of insulin in samples from M cows that had access to feed and water were greater (P < 0.001) than for VL cows (Figure 3B). Concentrations of insulin in fasted samples were greater (P < 0.001) for H and L cows than M cows, but VL cows were not different from H and M cows (Figure 3B). Concentrations of insulin were greater in samples from fed vs. fasted H (P < 0.001) and M (P < 0.001) cows, but insulin did not differ in samples from fed and fasted L and VL cows (Figure 3B). High cows had greater concentrations of leptin in fed (P < 0.001) and fasted (P < 0.001) samples at 109 d than cows on all other treatments (Figure 3C). Concentrations of leptin were greater in fed than fasted samples from H (P < 0.001) and M (P < 0.01) cows (Figure 3C). Low cows tended (P = 0.09) to have greater concentrations of leptin in fed samples than fasted samples, but concentrations did not differ in fed and fasted samples from VL cows (Figure 3C).
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Figure 3. Least squares mean concentrations of IGF-I (A, D), insulin (B, E), and leptin (C, F) in plasma of cows at 109 (A, B, C) and 123 d (D, E, F) of nutritional treatments: high (H = a 50% concentrate diet fed ad libitum in a drylot), or adequate native grass pastures and one of three amounts of a 40% CP supplement each day (M = moderate, 1.6 kg; L = low, 1.1 kg; or VL = very low, 0.5 kg; as-fed basis). a,b,cWithin fed or fasted cows, bars without a common letter differ, P < 0.05. y,zWithin a treatment, bars without a common letter differ, P < 0.05. Standard errors were 4.3, 0.06, and 0.4 for IGF-I, insulin, and leptin, respectively, on d 109, and 2.0, 0.06, and 0.3 for IGF-I, insulin, and leptin, respectively, on d 123 of treatment.
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On 123 d of the experiment, after cows had received the same diet for 13 d, BCS was greatest (P < 0.01) for H cows (6.4 ± 0.1), similar for M and L cows (4.8 ± 0.1), and least (P < 0.05) for VL cows (4.5 ± 0.1). Plasma concentrations of IGF-I (Figure 3D
), insulin (Figure 3E), leptin (Figure 3F), and thyroxine (Figure 2B) at 123 d were not influenced by previous nutritional treatments, and there was no interaction between treatment and access to feed. Concentrations of IGF-I (P < 0.001; Figure 3D) and insulin (P < 0.05; Figure 3E) were greater at 123 d in samples from fasted than fed cows (IGF-I = 18 ± 1 vs. 14 ± 1 ng/mL; insulin = 0.80 ± 0.02 vs. 0.73 ± 0.02 ng/mL, for fasted and fed cows, respectively). Plasma concentrations of leptin (Figure 3F) and thyroxine (Figure 2B) were not influenced by access to feed at 123 d.
Concentrations of IGF-I at d 68 were positively correlated (P < 0.01) with insulin in the plasma of cows that were fasted but not in cows that were fed (Table 1). Concentrations of leptin in plasma were not correlated with any of the hormones measured in plasma of fasted cows, and were positively correlated (P < 0.05) only with insulin in fed cows (Table 1). Thyroxine was positively correlated with IGF-I (P < 0.05) and insulin (P < 0.05) in plasma of fed cows (Table 1).
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Table 1. Correlation coefficients among insulin-like growth factor-I, insulin, leptin, and thyroxine in plasma of cows on 68, 109, and 123 d of treatment
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Concentrations of IGF-I in plasma at 109 d of treatment were positively correlated with insulin (P < 0.001) and leptin (P < 0.001) in samples from fed cows (Table 1). Insulin-like growth factor-I was also positively correlated with insulin (P < 0.001) and leptin (P < 0.05) after 18 h of fasting (Table 1). Concentrations of leptin in plasma at 109 d of treatment were positively correlated with insulin (P < 0.01) in fed but in not fasted samples (Table 1).
Concentrations of IGF-I in plasma samples from fed cows were not correlated with any of the hormones measured at 123 d; however, in samples from fasted cows, concentrations of IGF-I were correlated with insulin (P < 0.05) but not leptin (P = 0.35; Table 1). Concentrations of leptin were not correlated with insulin in either fed or fasted samples. However, concentrations of thyroxine were positively correlated with IGF-I (P < 0.01), insulin (P < 0.01), and leptin (P < 0.05) in samples from fasted cows (Table 1), and insulin was positively correlated with thyroxine (P < 0.05) in samples from fed cows (Table 1).
Effects of BCS
Body condition score of cows in this experiment ranged from 4.5 to 7.5 at 68 d, 4 to 7.5 at 109 d, and 3.5 to 7.5 at 123 d. Concentrations of insulin and leptin in samples from fed cows were correlated (P < 0.01; r = 0.63 and 0.36, respectively) with BCS at 68 d, and concentrations of IGF-I in plasma of fasted cows were correlated (r = 0.35; P < 0.01) with BCS. Body condition score at 109 d was positively correlated (P < 0.05) with plasma concentrations of IGF-I, insulin, and leptin in both fed (r = 0.55, 0.58, and 0.81, respectively) and fasted (r = 0.43, 0.30, 0.66, respectively) cows. Concentrations of IGF-I in samples from fed cows at 123 d, after all cows were on the same treatment for 13 d, were correlated (r = 0.27; P < 0.05) with BCS, and concentrations of leptin in both fed and fasted samples were correlated (P < 0.05; r = 0.28 and 0.35, respectively) with BCS. Concentrations of insulin and thyroxine were not correlated with BCS in samples from fed or fasted cows on d 123.
At 68 d of treatment, BCS accounted for 6% of the variation (P = 0.06) in concentrations of IGF-I in fasted samples, and 7% (P = 0.13; Table 2) of the variation in samples from fed cows (Figure 4A). There was a curvilinear relationship between BCS and concentrations of insulin in plasma, and BCS accounted for 47 (P < 0.001) and 31% (P < 0.001) of the variation in insulin from fed and fasted samples, respectively (Figure 4B). There was a linear relationship between BCS and leptin in plasma of fed and fasted cows at 68 d (R2 = 0.13; P < 0.01, and R2 = 0.25; P < 0.001, respectively; Figure 4C), and BCS accounted for 7% of variation in concentrations of thyroxine in fed cows (P < 0.05; Figure 5A) and was not signifi-cant for the variation in fasted cows.
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Table 2. Order of regression, R2, and P-value of the model when body condition score was regressed on concentrations of in insulin-like growth factor-I, insulin, leptin, and thyroxine in plasma of cows at 68, 109, and 123 d of treatment
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Figure 4. Relationships between concentrations of IGF-I (A), insulin (B), and leptin (C) in plasma with BCS of cows after 68 d of nutritional treatments: high (H = a 50% concentrate diet fed ad libitum in a drylot), or adequate native grass pastures and one of three amounts of a 40% CP supplement each day (M = moderate, 1.6 kg; L = low, 1.1 kg; or VL = very low, 0.5 kg; as-fed basis). Regression lines for insulin in fed and fasted cows differed, P < 0.05. Standard errors for IGF-I, insulin, and leptin were 2.6, 0.05, and 0.3, respectively.
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Figure 5. Relationships between concentrations of thyroxine in plasma and BCS of cows at 68 d (A) and 123 d (B) of treatment. Standard errors for thyroxine on 68 d and 123 d of treatment were 1.0 and 1.1, respectively.
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After 109 d of treatment, there was a positive curvilinear relationship between BCS and IGF-I (P < 0.001; Figure 6A) and leptin (P < 0.001; Figure 6C) in plasma of fed and fasted cows. Body condition score accounted for 34% (P < 0.001; Table 2) of the variation in concentrations of insulin in samples from fed cows, but only 10% (P < 0.05; Table 2) of the variation in samples from fasted cows (Figure 6B). At 123 d of the experiment, when all cows were on the same plane of nutrition, BCS accounted for less than 7% (Table 2) of the variation in concentrations of IGF-I and insulin, and BCS could not be used to predict concentrations of IGF-I (Figure 7A) or insulin (Figure 7B) in fed or fasted samples. There was a linear relationship between BCS and concentrations of leptin in samples from fed (P < 0.05) and fasted (P < 0.01) cows (Figure 7C). However, BCS only accounted for 7 and 12% of the variation in concentrations of leptin in fed and fasted cows, respectively. Concentrations of thyroxine in plasma of fed and fasted cows at 123 d (Figure 5B) were not strongly related to BCS.
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Figure 6. Relationships between concentrations of IGF-I (A), insulin (B), and leptin (C) in plasma with BCS of cows at 109 d of treatment. Regression lines for insulin (P < 0.05) and leptin (P < 0.001) in fed and fasted cows differed. Standard errors for IGF-I, insulin, and leptin were 2.8, 0.04, and 0.2, repectively.
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Figure 7. Relationships between concentrations of IGF-I (A), insulin (B), and leptin (C) in plasma with BCS of cows at 123 d of treatment. Regression lines for leptin in fed and fasted cows differed, P < 0.05. Standard errors for IGF-I, insulin, and leptin were 1.2, 0.03, and 0.2, respectively.
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Discussion
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Body fat reserves in cattle can be accurately predicted by BCS (Wagner et al., 1988; Yelich et al., 1995). Dietary treatments resulted in significant differences in BCS of cows. Body condition score was greatest for H cows, and VL cows consumed the least amount of supplemental protein and had the lowest BCS. Supplementation of cattle grazing low-quality forage with protein typically increases forage intake and BW gain (McCollum, III, and Horn, 1990).
Concentrations of insulin and leptin in plasma at 68 d, and concentrations of IGF-I, insulin, and leptin at 109 d were decreased with reduced nutrient intake. Nutrient restriction decreases plasma concentrations of IGF-I in cattle (Houseknecht et al., 1988; Rutter et al., 1989; Richards et al., 1995). Concentrations of insulin in cattle are decreased with restricted nutrient intake (Trenkle, 1978; Richards et al., 1989b; Bossis et al., 1999) and are associated with decreased concentrations of glucose in plasma (Richards et al., 1989b; Bossis et al., 1999). Concentrations of IGF-I at 109 d and insulin at 68 and 109 d, were less in H and M cows following 18 h of fasting, which agrees with previous reports for beef heifers (McCann and Hansel, 1986; Spicer et al., 1991). Body condition score was correlated with concentrations of IGF-I and insulin in plasma when cows had different amounts of nutrient intakes, but not when all cows received the same diet, suggesting that concentrations of IGF-I and insulin in plasma are indicative of metabolic status rather than the quantity of body fat.
Quantity of body fat is positively correlated with concentrations of leptin in pigs (Bidwell et al., 1997), and sheep (Blache et al., 2000). Concentrations of leptin in plasma of cows in the present experiment were positively correlated with BCS, a critical measure of body fat in the bovine. This finding agrees with a previous report for beef cows (Chilliard et al., 1998) when leptin was measured using a multispecies assay. Concentrations of leptin in plasma of Holstein cows at parturition and early lactation were greater in cows with more BCS (Meikle et al., 2004).
Increased nutrient intake was associated with increased BCS and increased concentrations of leptin in plasma. To separate the effects of nutrition from BCS, we evaluated concentrations of leptin in plasma from fed and fasted cows when they had different nutrient intakes and again after all cows were on the same diet. Previous nutritional treatment did not influence concentrations of leptin in plasma after all cows consumed the same diet for 13 d. This is in agreement with our previous report for postpartum primiparous cows, that feeding greater amounts of nutrients after parturition increased BCS and concentrations of leptin in plasma compared with moderately fed controls (Ciccioli et al., 2003). When nutrient intake was decreased from high to moderate amounts for the postpartum cows, concentrations of leptin were similar within 7 d, even though BCS differed (Ciccioli et al., 2003). The greater range in BCS in the present study expands our previous results. In the current experiment, BCS ranged from 3.5 to 7.5, which is equivalent to 6 to 20% carcass fat (Wagner et al., 1988). When cows were on the same diet, BCS accounted for less than 25% of the variation in plasma leptin vs. when cows were on different diets.
Plasma concentrations of leptin and insulin in L and VL cows were not influenced when feed and water were withheld for 18 h. A longer period of nutrient deprivation might have caused decreased concentrations of leptin in these cows. Plasma leptin in heifers and mature cows was decreased with 48 to 60 h of fasting (Amstalden et al., 2000, 2002). Chelikani et al. (2004) reported that the effect of fasting on plasma leptin occurred within 6 h in lactating Holstein cows, but not until 24 h in nonlactating cows. The amount of leptin mRNA in fat was decreased when pigs were fasted for 3 d (Spurlock et al., 1998). However, the abundance of leptin mRNA in adipose tissue of pigs fed maintenance, or 23% below maintenance energy requirements for 28 d, was not different than in pigs with ad libitum access to feed, despite the fact that the submaintenance pigs lost 14.4% of their initial BW (Spurlock et al., 1998). These results indicate that in pigs, leptin synthesis and/or secretion is not affected by moderate reductions in nutrient availability for a prolonged time period. The quantity of body fat in L and VL cows decreased during nutritional treatment, which was associated with decreased concentrations of leptin in plasma. Leptin in plasma of L and VL cows may have been at minimal concentrations by d 68 of treatment. Decreased concentrations of leptin in H and M cows in response to transient nutrient deprivation may function to stimulate appetite. Similarly, Delavaud et al. (2002) found that concentrations of leptin in fat Holstein and Charolais cows were decreased with underfeeding but were unchanged when lean cows were underfed (Delavaud et al., 2002). Thus, short-term control of plasma concentrations of leptin in cattle may be regulated by the amount of nutrient intake.
Decreased concentrations of leptin in plasma of M, L, and VL cows may be attributable to decreased synthesis of leptin. Thomas et al. (2001) determined that abundance of leptin mRNA in adipocytes of ewes on reduced nutrient intake was decreased compared with fully fed controls. Leptin mRNA in adipose tissue was decreased 50% after parturition in lactating Holstein cows in negative energy balance (Block et al., 2001). Amount of mRNA for leptin was decreased more than 30% in heifers (Amstalden et al., 2000) and cows (Tsuchiya et al., 1998) fasted for 48 h.
Adipose cell size is proposed as an important regulator of concentrations of leptin in plasma (Barb et al., 2001). The amount of leptin mRNA is correlated with the size of human (Wabitsch et al., 1996) and porcine (Chen et al., 1997, 1998) adipocytes. There was a positive quadratic relationship between adipose cell size and leptin concentration in plasma of beef cows (Delavaud et al., 2002). In the current experiment, concentrations of leptin were reduced on d 109 when H and M cows were fasted for 18 h, even though the duration of the fast was probably not sufficient to cause a change in body fat mass or adipose cell size. Thus, changes in plasma leptin independent of changes in the amount of body fat indicate that leptin synthesis and/or secretion may be responsive to hormonal control rather than the size of fat cells alone in ruminants.
Insulin is a potent stimulant of adipocyte maturation and lipid filling (Chen et al., 1998). In the present experiment, leptin was correlated with IGF-I and insulin at d 109, and changes in plasma leptin paralleled changes in insulin and IGF-I. This was also the case with pigs (Spurlock et al., 1998) and fasted heifers (Amstalden et al., 2000). Insulin increased in vitro release of leptin from adipose tissue of rodents (Ardie et al., 1996) and increased the abundance of leptin mRNA in adipocytes of pigs (Chen et al., 1998). Messenger RNA for leptin was decreased with fasting in rats (Saladin et al., 1995) and mice (Mizuno et al., 1996), but treatment of fasted rodents with insulin restored amounts of leptin mRNA (Saladin et al., 1995; Mizuno et al., 1996), which occurred independently of concentrations of glucose in plasma (Saladin et al., 1995). Amounts of leptin mRNA in bovine adipose tissue explants were increased with insulin treatment (Houseknecht et al., 2000), indicating that leptin synthesis in the ruminant may be influenced by insulin. Delavaud et al. (2002) found that insulin was not correlated with concentrations of leptin in serum of cows that were underfed for 1 wk. In contrast, animals in the current study had different nutrient intakes for 3 mo, which may have potentiated effects of insulin on adipose tissue function.
Whether IGF-I is a major regulator of leptin in not clear. Concentrations of IGF-I and leptin in heifers fasted for 48 h were decreased (Amstalden et al., 2000). In the current experiment, IGF-I and leptin were positively correlated on d 109 of treatment. Abundance of mRNA for leptin and IGF-I in adipose tissue of steers was positively correlated (Houseknecht et al., 2000), indicating that local production of IGF-I may be associated with abundance of leptin mRNA. Insulin-like growth factor-I increased the amount of leptin mRNA abundance in differentiated porcine adipocytes (Chen et al., 1998). However, expression was less than when adipocytes were treated with insulin, indicating that insulin may be a more potent regulator of leptin gene expression than IGF-I in swine. In contrast, Hardie et al. (1996) found that IGF-I did not alter the abundance of leptin mRNA in rat adipocytes in vitro. Different effects of IGF-I on leptin mRNA abundance may reflect species differences in metabolism.
Hormonal and/or metabolic regulation of leptin may be mediated by factors other than insulin or IGF-I. Volatile fatty acids are produced by ruminal fermentation and are used as sources of energy. In well-fed cows, plasma concentrations of leptin were negatively correlated with concentrations of 3-OH-butyrate in blood 4 h after consumption of feed (Delavaud et al., 2002), and intravenous administration of propionate increased plasma concentrations of insulin and expression of leptin mRNA in adipose tissue in sheep (Lee and Hossner, 2002). The effects of nutritional treatments on plasma concentrations of VFA in the current experiment were not determined. Additionally, relative amounts of individual VFA in blood vary with composition of nutrient intake and may regulate expression of leptin mRNA indirectly via their effect on other constituents in plasma, such as insulin.
Cows on the greatest nutrient intake had greater concentrations of thyroxine in plasma at 68 d than cows on all other treatments, but thyroxine was not correlated with leptin. Concentrations of thyroxine and leptin were positively correlated in fasted samples at 123 d. We previously found that leptin and thyroxine concentrations in plasma of primiparous heifers were correlated (Ciccioli et al., 2003). Delavaud et al. (2002) found that decreased nutrient intake resulted in decreased concentrations of thyroxine and leptin in plasma of beef cows, but these two hormones were not correlated with each other. Different concentrations of thyroxine between treatments probably reflect differences in metabolic rate rather than a direct effect on leptin.
Nutrient intake influences concentrations of leptin in mature gestating beef cows. Plasma concentrations of leptin are correlated with concentrations of insulin and IGF-I in plasma when cows have different nutrient intakes and BCS. Although concentrations of leptin in plasma are correlated with BCS, nutritional effects can occur independently of the amount of body fat. Decreased circulating concentrations of leptin coincide with decreased concentrations of insulin and IGF-I. These results are similar to those for rodents and pigs, and indicate the potential for hormonal regulation of leptin synthesis and/or secretion in the ruminant. Additionally, nutrient deprivation for as little as 18 h is sufficient to decrease plasma concentrations of IGF-I, insulin, and leptin in the bovine during mid-gestation. Time of sample collection relative to availability of feed may influence results of nutritional or metabolic experiments in the bovine. Body condition score of cows significantly influenced concentrations of leptin, insulin, and IGF-I in plasma when cows had different nutrient intakes. However, when gestating cows had similar nutrient intakes, BCS only accounted for less than 12% of the variation in concentrations of leptin in plasma and did not influence plasma concentrations of insulin and IGF-I. The effects of nutrient intake of gestating beef cows on plasma concentrations of leptin, insulin, and IGF-I are greater than the effect of body fat reserves.
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Implications
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Concentrations of insulin, insulin-like growth factorI, and leptin in plasma of beef cows are indicators of the adequacy of nutrient intake but are not highly correlated with body condition score. Because body condition score or the amount of body fat is a major regulator of reproductive performance, plasma concentrations of insulin, insulin-like growth factor-I, and leptin may be inadequate as predictors of potential reproductive performance of beef cows. Additional research is necessary to elucidate how greater nutrient intake, and associated increases in concentrations of insulin, insulin-like growth factor-I, and leptin in plasma, may be used to enhance reproductive efficiency when body energy reserves are inadequate.
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Footnotes
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1 Approved for publication by the Director, Oklahoma Agric. Exp. Stn. This research was supported under Project H-2331. 
2 Current address: Animal Reproduction and Biotechnology Lab, Colorado State Univ., Fort Collins 80523.
4 Current address: Dept. of Vet. Anat. and Public Health, Texas A&M Univ., College Station 77843.
3 Correspondence: 114 Animal Science (phone: 405-744-6077; fax: 405-744-7390; e-mail: rpw{at}okstate.edu).
Received for publication August 4, 2004.
Accepted for publication November 19, 2004.
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Abstract
Introduction
