The Effect of Photoperiod on the Profile of Prolactin, Leptin, Insulin and the Content of Bioactive Substances in Sheep Milk During the Rearing Period of Lambs

TL;DR

Longer daylight hours during lambing increase milk production and lamb growth in sheep through hormonal and nutritional changes in milk.

Contribution

Demonstrates how photoperiod affects milk hormones, composition, and lamb growth in sheep, offering practical insights for sheep production.

Findings

Ewes lambing in longer daylight periods produced milk with higher prolactin and insulin levels and higher milk yield.

Lambs from long-day ewes had greater daily weight gains compared to those from short-day ewes.

Milk from short-day ewes had higher dry matter, fat, protein, and lactose content.

Abstract

Seasonal changes in day length strongly influence physiological processes in sheep, including milk production and lamb growth. This study examined how the season of lambing affects milk hormone levels (prolactin, leptin, and insulin), the nutrient composition of milk, and lamb body weight gain. Ewes that lambed under longer daylight hours produced more milk with higher prolactin and insulin levels, and their lambs showed greater daily body weight gains compared to mothers lambing under shorter daylight hours. Differences in milk nutrient content were also found between seasons. These studies demonstrate that seasonal environmental factors can alter both milk quantity and quality, thus influencing offspring growth. Understanding these effects helps improve sheep management practices, optimal planning of production seasons, and strategies that support both milk yield and lamb development, which is important for animal welfare and animal production efficiency.

Photoperiod and seasonality influence reproduction and lactation in sheep, but their effects on milk hormones, milk composition, and lamb growth are not fully understood. This study assessed the effect of season on milk prolactin, leptin, and insulin concentrations, milk chemical composition, lactation performance, and lamb growth in Polish Mountain ewes. Forty ewes were divided into the following two groups: short-day (lambing in December, n = 20) and long-day (lambing in May, n = 20). Milk samples were collected on days 5, 15, 25, 35, and 45 of lactation. Ewes in the long-day photoperiod had higher milk yield (p < 0.01) and higher prolactin and insulin concentrations (p < 0.01), whereas leptin concentrations did not differ seasonally. Milk from short-day ewes was characterized by higher dry matter and fat content (p < 0.01) and higher protein and lactose content (p < 0.05). Lambs from the long-day group achieved higher mean daily gain (p < 0.01). These results indicate that photoperiod influences lactation performance, milk composition, and offspring growth through seasonal hormonal and metabolic mechanisms, suggesting that appropriate lambing timing and day length manipulation can improve milk production efficiency and lamb growth in practical sheep production systems.

Full text

1. Introduction

Sheep (Ovis aries) exhibit distinct seasonal changes in reproductive activity, dependent on day length (photoperiod). In most sheep breeds, photoperiod acts as a key regulator synchronizing annual reproductive cycles [1]. Light received by retinal photoreceptors influences the central nervous system and leads to melatonin secretion by the pineal gland [2]. In sheep, as a “short-day” species, longer nights (i.e., shortening photoperiod) are associated with increased melatonin secretion, which stimulates the hypothalamic–pituitary–gonadal axis (HPG), promoting gonadotropin release and supporting ovarian follicle development and the onset of estrus [1,2,3]. This neuroendocrine mechanism underlies seasonal breeding in sheep.

Seasonal breeding also influences the lactation period. In addition to melatonin, prolactin (PRL) plays a key role in regulating mammary gland function and milk production [4]. Changes in photoperiod influence the rhythm of PRL secretion, which translates into mammary gland activity and milk yield [4]. During periods of short daylight, when melatonin is more abundant, PRL secretion is reduced, which may limit milk production [5,6]. The lambing season, meaning the day length at the onset of lactation, can influence total milk yield. Previous studies have shown that ewes lambing during lengthening days had longer lactations and higher milk production than those that began lactation during shortening days [7,8]. Consequently, seasonality of reproduction is closely linked to seasonality of lactation, and changes in photoperiod and the associated hormonal fluctuations determine both fertility and milk yield in sheep.

In addition to classic photoperiod hormones such as melatonin and prolactin, there is a growing interest in the role of metabolic hormones, particularly leptin (LEP) and insulin, in regulating mammary gland function and milk composition. LEP, primarily known as a regulator of appetite and energy balance, can also influence endocrine pathways related to metabolism and growth, and its effects may vary depending on day length and seasonal tissue sensitivity in sheep [9,10]. Leptin is also present in milk and mammary gland tissues, where it may perform paracrine and endocrine functions [11]. Recent studies have shown that leptin signaling in mammary epithelial cells can activate the PI3K/AKT/mTOR pathway, which increases the expression of genes involved in milk fat synthesis, providing evidence for a direct role of leptin in the metabolism of milk components [12]. Furthermore, in vitro models have observed that leptin can modulate the activity of transcription factors involved in milk protein synthesis, including leptin, by inhibiting STAT5 in mammary epithelial cells, which further indicates its complex role in the regulation of lactation [13]. Insulin regulates glucose metabolism and acts as a growth factor both before and after weaning, influencing nutrient transport and endocrine regulation in the offspring [14,15]. Recent studies confirm its key role in regulating glucose metabolism and nutrient utilization in the mammary gland, which influences lactose and milk protein synthesis and is therefore essential for maintaining high lactation efficiency and milk quality. By activating PI3K-Akt signaling and related metabolic pathways, insulin supports the activity of mammary epithelial cells and promotes anabolic processes associated with milk production [16]. Available data also demonstrate the presence of insulin in milk and the variability of its concentrations depending on the stage of lactation, highlighting its biological importance in mammary gland metabolism [17].

It is proposed that interactions between PRL, LEP, and insulin—in the context of photoperiod and seasonality—create an adaptive regulatory network that integrates signals about energy status, environmental conditions, and reproductive needs. Such a network may enable animals to optimize reproduction, lactation, and offspring development in a changing environment [10]. Available data indicate that, in addition to classic photoperiod hormones (melatonin, gonadotropins), metabolic hormones such as LEP and insulin may also play a significant role in regulating lactation, milk production, and offspring development in sheep [9]. PRL remains the key hormone activating the mammary gland, while LEP and insulin transmit metabolic signals [18]. The interaction of these hormones, which are subject to seasonal and metabolic changes, can modulate both the chemical composition of milk, its quantity, and the growth rate of lambs [19].

The chemical composition of milk is a central element in assessing the quality of lactation, directly influencing its nutritional value and biological role in offspring nutrition. Sheep milk is characterized by a relatively high content of dry matter, fat, protein, and lactose, which distinguishes it from the milk of other species and contributes to its energy-rich nutritional profile [20]. The composition of these components is determined by numerous factors, including species physiology, season, photoperiod, nutritional conditions, and female health status, and therefore changes with environmental conditions and the stage of lactation.

Therefore, the aim of this study was to determine the effect of the season on the levels of PRL, LEP and insulin in sheep milk, the chemical composition of milk and the daily weight gain of lambs during the lamb rearing period.

2. Materials and Methods

2.1. Animals and Management

The Polish Mountain Sheep is a native, locally preserved breed of sheep with a moderate utility profile, well-adapted to the harsh environmental conditions of the mountains and foothills. This breed has an open, wooly coat, a muscular build, and is characterized by a relatively high prolificacy of approximately 150%, with ewes weighing an average of 45–55 kg and rams 60–70 kg, reflecting its reproductive potential compared to other native sheep breeds in Poland. The Polish Mountain Sheep has been the subject of genetic resource conservation programs and is characterized by the seasonality of breeding typical of mountain breeds and a moderate lambing rate, making it a valuable element of the genetic diversity of Polish sheep farming [21]. The animals were matched according to age (3–4 lactations) and body weight (45–50 kg). They were farmed at the Experimental Station of the Faculty of Animal Sciences of the University of Agriculture in Krakow (Poland) under natural lighting conditions (50°04′ N, 19°57′ E). During the short-day period, the length of the day was 8 h of light and 16 h of darkness, while during the long-day period, it was 16 h of light and 8 h of darkness. The animals were fed according to the physiological status based on the standards of the National Research Institute of Animal Production (Krakow-Balice, Poland) [22].

From the beginning of the experiment (mating) to 4 months of pregnancy, sheep were fed forage pasture 5 kg/per sheep/day (dry matter: 214 g, crude protein: 49 g, and net energy: 1.24 MJ, per kg), silage 4 kg/per sheep/day (dry matter: 382 g, crude protein: 58 g, and net energy: 1.95 MJ, per kg) and hay ad libitum (dry matter: 882 g, crude protein: 185 g, and net energy: 3.24 MJ, per kg). The sheep had unlimited access to water and salt licks.

In order to limit the influence of the nutritional factor on the chemical composition of milk, the nutrition was unified from the 4th month of pregnancy to the end of the experiment. Accordingly, sheep received 1.0 kg of pelleted granulate complete feed (crude protein: 220 g and net energy: 7.5 MJ, per kg containing only natural components—cereal grains, rape, dried legume plants, dried beet pulp, and corn flour) produced under the name CJ by Polish company, and hay supplement (dry matter: 882 g, crude protein: 185 g, and net energy: 3.24 MJ, per kg).

The total daily requirement was divided into two portions, morning (500 g CJ, hay ad libitum/per sheep) and evening (500 g CJ, hay ad libitum/per sheep). The hay for experimental animals was collected in the first cut before flowering at the same time from the same meadow of the Experimental Station of the University of Agriculture. All animals had free access to water and mineral licks. The zoohygienic and weather conditions were the same for all animals and body condition score was 3 or 4 [23].

2.2. Experimental Design and Sample Collection

Sheep were divided into two treatment groups of 20 animals each (G1 n = 20 and G2 = 20). In each group, estrus synchronization was performed in a different photoperiodic period using the Chronogest method. Specifically, polyurethane sponges impregnated with fluorogestone acetate (Chronogest^®^) were inserted intravaginally for 14 days to mimic the luteal phase by providing a sustained progestagen exposure, which suppresses natural estrus behavior and ovulation during treatment. Upon sponge removal, ewes received an intramuscular injection of 500 IU pregnant mare’s serum gonadotropin (PMSG) to stimulate synchronized follicular development and ovulation. Estrus typically occurred within 24–48 h after PMSG administration, and timing was confirmed using a teaser ram. This protocol is a well-established approach in sheep reproductive management for inducing and synchronizing estrus [24]. Mating after estrus synchronization was performed from the 1st to 5th July (G1) and from the 1st to 5th December (G2). Therefore, sheep from group G1 lambed in December (short-day group), and those from group G2 lambed in May (long-day group). Both groups of sheep were maintained together with their lambs under natural daylength conditions. Lambs stayed with their mothers until 56 days of age. In both groups, individual milk samples were taken on the 5th, 15th, 25th, 35th and 45th day. For the sampling of milk, the sheep were hand milked at 8:00 a.m. Cumulative samples of milk were made on the right and left halves of the udder. There were 80 samples in the season 1 and 80 samples in the season 2. After collecting the milk samples, they were frozen at the temperature of −20 °C until the analysis (14 days). All the sheep in the experiment fed the twins.

2.3. Hormone Concentrations in Milk

The concentration of PRL in milk was assayed by the radioimmunoassay (RIA) double-antibody method using anti-ovine prolactin NIDDK-anti-oPRL-2 (AFP-C358106, HUMC, Torrance, CA, USA) and anti-rabbit gamma globulin antisera R0881-5000TST (Sigma-Aldrich, Saint Louis, MO, USA) [25]. The milk samples were diluted 10-fold, the range of the calibration curve was from 3 to 400 ng/mL, and the working dilution of anti-ovine prolactin antiserum was 1:20,000. The assay sensitivity of the assay was 2 ng/mL, and the intra- and interassay coefficients of variation were 9 and 12%, respectively. Milk LEP concentration was measured using a Multi-Species leptin RIA kit (Cat. No. XL-85K, Sigma-Aldrich, Saint Louis, MO, USA), according to the manufacturer’s instruction. The range of the calibration curve was from 1 to 50 ng/mL; the assay sensitivity was 1 ng/mL, and the intra- and interassay coefficients of variation were 3.6 and 8.7%, respectively. Quantitative determination of insulin concentration in sheep’s milk was performed using a Mercodia Ovine Insulin ELISA kit (Cat. No. 10-1202-01, Mercodia AB, Uppsala, Sweden), according to the manufacturer’s instruction. The range of the calibration curve was from 0.05 to 3.0 µg/L; the assay sensitivity was 0.025 µg/L, and the intra- and interassay coefficients of variation were 3.7 and 6.8%, respectively. Before the assays, samples of thawed milk were sonicated for 10 min and skimmed by centrifugation (4000× g for 15 min).

2.4. Chemical Compositions

The chemical composition of milk was determined using standardized analytical methods [26]. Dry matter content was assessed by the gravimetric drying method at 102 °C until constant weight, according to PN-ISO 6731:2014-11P [27] and AOAC [28] official procedures for moisture and solids analysis. Total protein content was measured by the Kjeldahl method, which quantifies total nitrogen that is then converted to crude protein using an appropriate conversion factor, following the procedure described in PN-68/A-86122 [29]. Fat content was determined by the Gerber butyrometric method, a reference procedure for dairy fat analysis specified in PN-68/A-86122 [29]. Lactose was quantified using the Bertrand titrimetric method as described in PN-68/A-86122 [29]. Calcium and phosphorus concentrations were determined by atomic absorption spectrometry (AAS) using an AA240 FS spectrometer (Varian Inc., Mulgrave, Australia), in accordance with AOAC spectrometric protocols. The dry matter content was analyzed using dryer method [PN-ISO 6731:2014-11P] [27] [AOAC 1995] [28] and protein content using methods by Kjeldahl [PN-68/A-86122] [29]. The fat content was analyzed according to the methods of Gerber [PN-68/A-86122] [29]. The lactose was analyzed using the methods Bertrand [PN-68/A-86122] [29]. Calcium and phosphorus concentrations were determined by flame atomic absorption spectrometry (AAS) using an AA240 FS spectrometer, according to AOAC methods [28]. Fatty acids were determined with gas chromatography (chromatograph PYE-UNICAM series 104 with chromatography column SUPELCOWAX 10.30 m, ø 0.53 mm, 1.0 µm) [30]. Separation conditions were as follows: helium carrier gas 2.5 mL / min; dispenser temperature 235 °C; oven temperature 50 °C for 2 min; 7 °C for 1 min up to 210 °C; 210 °C for 20 min; and detector temperature 250 °C (flame-ionization FID). A purified preparation of DEGS, 1 and attenuation at 32 was used. The amino acid profile (asp, ser, glu, gly, his, arg, thr, ala, prol, cys, tyr, val, met, lys, ile, leu, and phe) was determined by reverse-phase liquid chromatography using an analytical kit ACCQ Tag (Waters, Milford, MA, USA). Hydrolysis of about 30 mg of the sample was carried out with 4 mL of 6M HCl (POCH, Gliwice, Poland) and the addition of 15 μL of phenol (Sigma-Aldrich, St. Louis, MO, USA) at the temperature of 110 °C for 24 h. The obtained hydrolyzate was filtered through 0.45 µm syringe filters and then dried with nitrogen. The sample prepared in this way, after appropriate dilution, was subjected to the derivative procedure according to the Waters company recommendations. In the case of standards (Waters, Milford, MA, USA), the procedure was analogous. The chromatographic separation was performed using a liquid chromatograph by Thermo Scientyfic: Dionex Ultimate 3000 equipped with a 4-channel gradient LPG pump—3400 SD, WPS 3000 TSL autosampler and a 4-channel FLD-3400RS fluorescence detector. A Nova-Pak C 18.4 µm column (150 × 3.9 mm) from Waters USA was used for the analysis. Separation temperature 37 °C. Elution was performed in a 1 mL/min binary gradient as follows: eluent A acetate-phosphate buffer, B acetonitrile/water 60:40, according to the manufacturer’s standard Waters procedure. Quantitative analysis was performed using a 1-point calibration (using an analytical standard of 100 pmol each). The results were developed using Chromeleon 7.0 software.

2.5. Milk Yield and Daily Growth of Lambs

Lambs were weighed twice—on days 2 and 28 of life—to estimate body weight gain in the first postpartum period. A conversion factor of 4.5 L of ewes’ milk for every 1 kg increase in lamb body weight was used to correct the growth data. This allowed for linking growth rate with the ewes’ lactation efficiency in the pre-weaning period. After 28 days of age, the lambs began supplementing with solid feed, which changed the energy and protein sources available to young animals. Consequently, after 28 days of age, daily gains of lambs were no longer related to the mother’s milk yield, and subsequent growth rate was analyzed independently of lactation parameters, due to the significant contribution of supplementary feeding to the lambs’ nutritional balance.

2.6. Statistical Analysis

The hormone concentrations in milk and chemical compositions data were analyzed using the two-way repeated-measures analysis of variance (ANOVA, Statistica; StatSoft, Inc., Tulsa, OK, USA). After the ANOVA, the Tukey post hoc test was performed, when appropriate. The Kruskal–Wallis test followed by multiple comparisons of average ranks (Statistica; StatSoft, Inc., Tulsa, OK, USA) was used to determine the significance of the differences in chemical compositions among the groups. Differences were considered significant at p < 0.05 with tendencies discussed at p ≤ 0.05 ≤ p ≤ 0.01. All data are expressed as means ± SEM.

3. Results

3.1. Milk Concentrations of Prolactin, Leptin and Insulin

As shown in Table 1, PRL concentration in sheep’s milk differed significantly between groups G1 (short-day photoperiod) and G2 (long-day photoperiod) throughout lactation. On day 5, ewes from group G2 showed significantly higher PRL levels (277.92 ng/mL ± 21.4) compared with group G1 (58.05 ng/mL ± 9.84; p < 0.01). Similar patterns were observed on subsequent sampling days as follows: on day 15, PRL concentration was 195.59 ng/mL ± 20.6 in group G2 compared with 39.45 ng/mL ± 8.33 in G1 (p < 0.01); on day 25: 189.57 ng/mL ± 13.7 vs. 92.39 ng/mL ± 11.19 (p < 0.01); on day 35: 230.82 ng/mL ± 19.8 vs. 63.17 ng/mL ± 9.94 (p < 0.01); and on day 45: 249.72 ng/mL ± 20.9 vs. 59.17 ng/mL ± 9.94 (p < 0.01). Total mean PRL concentration at all time points was significantly higher in group G2 (228.72 ng/mL ± 18.79) than in group G1 (62.45 ng/mL ± 9.84; p < 0.01). These results indicate a consistent and statistically significant difference in PRL secretion between the two photoperiod groups in early lactation.

As shown in Table 2, LEP concentration in sheep milk did not differ significantly between group G1 (short-day photoperiod) and group G2 (long-day photoperiod) on any of the sampling days examined. On day 5 of lactation, the mean LEP concentration was 25.18 ng/mL (±8.45) in group G1 and 27.97 ng/mL (±8.11) in group G2 (NS). Similar non-significant differences were observed on day 15 (G1: 23.56 ng/mL ± 7.13; G2: 27.28 ng/mL ± 7.98, NS), day 25 (G1: 23.68 ng/mL ± 6.98; G2: 19.05 ng/mL ± 5.28, NS), day 35 (G1: 24.74 ng/mL ± 7.21; G2: 24.32 ng/mL ± 6.72, NS) and day 45 (G1: 23.64 ng/mL ± 6.28; G2: 25.22 ng/mL ± 6.77, NS). The overall mean LEP concentration across all sampling points was 24.14 ng/mL (±6.58) in G1 and 24.76 ng/mL (±6.87) in G2, with no statistically significant effect of photoperiod on milk LEP content (NS).

As shown in Table 3, insulin concentrations in sheep’s milk differed significantly between the two photoperiod groups throughout the lactation period. On day 5, ewes in group G2 (long-day photoperiod) had an insulin concentration of 0.50 µg/L (±0.03), which was significantly higher than 0.25 µg/L (±0.01) observed in group G1 (short-day photoperiod) (p < 0.01). Similar significant differences were noted at day 15 (G1: 0.24 ± 0.01 vs. G2: 0.55 ± 0.03, p < 0.01), day 25 (G1: 0.31 ± 0.02 vs. G2: 0.54 ± 0.03, p < 0.01), day 35 (G1: 0.39 ± 0.02 vs. G2: 1.06 ± 0.03, p < 0.01) and day 45 (G1: 0.42 ± 0.02 vs. G2: 1.03 ± 0.03, p < 0.01). The overall mean insulin concentration over the entire 45-day period was significantly higher in group G2 (0.73 ± 0.03 µg/L) than in group G1 (0.32 ± 0.01 µg/L; p < 0.01).

In group G1 (sheep rearing lambs during the short-day period), the highest prolactin concentrations in milk were observed on days 25 and 35 of lactation, with the lowest values observed on day 15. The mean prolactin level during the first 45 days of lactation was 62.45 ng/mL. Leptin showed little fluctuation over time, with the lowest level on day 15 and a mean of 24.14 ng/mL. Insulin in group G1 reached its lowest levels on day 15, with a mean concentration of 0.32 µg/L, with a significant increase in the later days of lactation.

In group G2 (sheep rearing lambs during the long-day period), prolactin was significantly higher than in group G1 throughout the study, with a mean of 228.72 ng/mL and the highest values at the beginning of lactation. The mean leptin concentration was 24.76 ng/mL and did not differ significantly between days. Insulin in G2 was significantly higher than in G1 at all measurement points, with the highest values on days 35 and 45 of lactation and a mean of 0.73 µg/L.

3.2. Chemical Composition of Milk

As shown in Table 4, significant differences were observed in the chemical composition of sheep’s milk between the two groups. Ewes lambing in December (G1) produced milk with a significantly higher dry matter content (16.23 ± 5.20%) compared to ewes lambing in May (G2; 13.74 ± 4.90%; p < 0.01). Protein content was also significantly higher in G1 (5.74 ± 1.30%) than in G2 (5.66 ± 1.20%; p < 0.05). Similarly, fat levels were significantly higher in G1 (4.69 ± 0.50%) than in G2 (3.65 ± 0.80%; p < 0.01). Moreover, lactose content was significantly higher in milk from G1 sheep (4.82 ± 0.70%) compared to G2 (4.50 ± 0.80%; p < 0.05).

3.3. Milk Yield and Daily Gains of Lambs

As shown in Table 5, milk yield during the first 28 days of lactation was significantly different between the two groups. Ewes in group G2 (long-day photoperiod) produced significantly more milk (41.4 ± 5.6 L) than ewes in group G1 (short-day photoperiod; 24.4 ± 4.2 L; p < 0.01). Furthermore, the mean daily body weight gain of lambs was significantly greater in group G2 (310 g/day ± 11.6) compared with lambs in group G1 (240 g/day ± 510.1; p < 0.01).

4. Discussion

The study demonstrated that lambing season and associated photoperiod differences influence hormone concentrations in milk, milk chemical composition, lactation performance, and offspring growth rate in ewes. Hormone profile analysis revealed significant differences in PRL and insulin levels in milk between groups of ewes rearing lambs under short- and long-day photoperiod conditions, consistent with the concept of a seasonal mechanism for regulating lactation in ruminants. In contrast, LEP levels in milk did not show significant seasonal differences. Significantly higher PRL concentrations were found in the milk of ewes rearing lambs under long-day conditions (group G2) on all days of lactation compared to the milk of ewes rearing lambs under short-day conditions (group G1). This indicates that photoperiod, through its effect on the neuroendocrine axis and melatonin secretion, modulates PRL secretion in sheep, which translates into lactation intensity and hormonal adaptation to the season [8,31]. Similarly, the literature describes seasonal rhythms of PRL in sheep, with its concentration reaching higher values in summer (long daylight) than in winter (short daylight), which influences milk production and lactation parameters [6].

Unlike PRL, LEP in milk did not differ significantly between groups. The lack of significant seasonal differences in milk may be due to the fact that its concentration in milk is more closely related to the overall metabolic balance and energy status of the ewe than directly to photoperiod, as suggested by studies on LEP in the serum and tissues of seasonal mammals [6]. Its role in milk is biologically important, as it participates in gastrointestinal development and offspring metabolism. However, seasonal modulation of its secretion into milk may be more subtle and dependent on the metabolic status of the ewes themselves.

Studies have shown that milk insulin concentrations were significantly higher in sheep in the G2 phase on all days of lactation. Although the literature on seasonal variation in insulin in ruminant milk is still limited, it has been suggested that photoperiod may also modulate the transport and secretion of metabolic hormones into milk, supporting glucose metabolism and offspring tissue growth. Similar relationships have been observed in studies of sheep and other seasonal species, where photoperiod influenced the levels of metabolic hormones such as insulin-like growth factor (IGF-1) and insulin, particularly under long-day conditions [32]. Insulin in milk may act as a bioactive metabolic signal, supporting glucose metabolism, nutrient transport, and anabolic processes in the mammary gland and offspring, which is important for milk protein and lipid synthesis and young animal growth. Insulin-like growth factor IGF-1, present in milk, participates in the proliferation of mammary epithelial cells and the synthesis of milk components, and its signaling supports the development and functioning of the mammary gland even during lactation [33]. Furthermore, IGF-1 present in milk has been associated with offspring growth parameters, such as body weight gain and developmental indices, in other studies, suggesting its role as a bioactive growth factor for newborns [34]. In the context of photoperiod, higher insulin concentrations with longer days may reflect the adaptive metabolic adjustment of the body to more intensive lactation and the energy needs of the offspring [35].

Analysis of the chemical composition of milk revealed clear seasonal differences between the studied groups of sheep. Milk from ewes raising lambs under short photoperiod conditions (group G1) was characterized by significantly higher dry matter, protein, fat, and lactose contents compared to milk from G2 mothers raised under longer-day conditions. The increased dry matter content in G1 milk may have contributed to the simultaneous increase in concentrations of other key chemical components, indicating the possible influence of seasonal conditions and maternal metabolism on the milk profile during the short-day period. High dry matter content is a condition favoring increased protein and fat concentration, and has been previously confirmed in the studies by Molik et al. (2011; 2020) [8,31], which showed that sheep milk with higher dry matter content is also characterized by increased amounts of these components. In the context of the analyzed results, the significantly higher protein content in the milk of G1 ewes may reflect both seasonal fluctuations in the regulation of milk protein synthesis and potential similarity to values observed in Manchega ewes under short-day conditions [36], which is also confirmed by earlier observations by Molik et al. (2011) [31]. Similarly, the increased fat content in the milk of G1 ewes indicates a change in the lipid profile in response to a shorter photoperiod, which is consistent with the results of studies on seasonal variation in fat fractions in sheep milk [31,37]. In the case of lactose, its content in the milk of G2 ewes was significantly lower than in the milk of G1 ewes, which also aligns with previously observed seasonal differences in this nutrient concentration [8], and lactose values in G1 milk are similar to those described in Manchega and Lacaune ewes [36]. Overall, the results indicate that photoperiod and related hormonal rhythms can modulate the chemical profile of ewes’ milk and its potential to meet the energy and structural needs of offspring [8]. The higher protein and fat concentrations in the milk of G1 ewes may indicate an adaptive metabolic strategy during shorter days, when environmental conditions are less favorable, leading to physiological changes in the mechanisms of milk component synthesis.

Physiologically, photoperiod modulates the secretion of key hormones associated with lactation, which may partially explain the observed differences in milk yield and milk composition between groups. In seasonally responsive ewes, shorter daylight hours lead to increased melatonin secretion, which in turn inhibits prolactin secretion (a hormone central to the synthesis and maintenance of lactation) and may limit the milk production rate observed in the short-day group. Conversely, longer days promote higher prolactin concentrations and are associated with increased milk production, whereas under short photoperiods, reduced prolactin concentrations are associated with reduced lactation performance in ewes. The literature describes seasonal prolactin suppression and decreased milk production under shorter daylight conditions [6,8]. These mechanisms involve melatonin acting as a photoperiod signal on the HPG, resulting in changes in the hormonal profile and metabolic adaptations of the mammary gland and the entire organism. In response to lower milk yield under short-day conditions, the organism may compensatory increase the concentration of milk components, reflecting the observed higher chemical composition of milk with a concomitant lower volume, as suggested by studies on photoperiod and melatonin in sheep [8].

It should be emphasized, however, that the chemical composition of sheep’s milk in both analyzed groups could also have been influenced by other factors. In addition to the variable photoperiod, the presented results could have been influenced by characteristics such as genetic differences between individuals and breeds, herd management, environmental conditions (temperature, humidity), farm location, and udder health [38]. Particular attention should be paid to the animals’ diet, its quality, and availability. During the experiment, the sheep received the same amount of pelleted complete feed and ad libitum hay from the same harvest batch, aimed at standardizing feeding conditions between the groups. Nevertheless, it cannot be ruled out that, for example, differences in the amount of hay consumed could have influenced some of the observed differences in the chemical composition of the milk. Furthermore, lamb gender was not considered as a factor in the analysis of body weight gain, which may constitute an additional limitation in the interpretation of the results. In addition, ambient temperature was neither recorded nor controlled throughout the study period, and variable thermal conditions may have influenced not only lamb growth and body weight gain, but also ewe physiology and milk production, thereby potentially affecting the overall results.

A significant finding of the analysis was that ewes’ milk yield and lamb growth rate differed significantly between groups. Ewes from group G2 produced more milk than ewes from group G1, and lambs from group G2 achieved higher average daily weight gains. These results indicate that photoperiod influences both the amount of milk produced and the efficiency of its utilization by the offspring. Higher milk yield may have increased the availability of nutrients necessary for lamb growth, which was reflected in their faster growth rates. Similar seasonal relationships between milk yield and offspring growth have been reported previously, with shorter days associated with lower milk production and longer days with increased milk production [7,39]. Such seasonal differences suggest that light conditions and photoperiod-related hormonal rhythms may modulate both milk production mechanisms in ewes and milk utilization by lambs.

From a biological perspective, higher concentrations of hormones such as PRL and insulin, as well as increased milk production, may support lamb metabolism and growth by increasing energy availability and modulating metabolic signals, as reflected in the higher daily weight gains observed in the G2 group. At the same time, changes in milk chemistry may represent an adaptive response to seasonal environmental and metabolic conditions, allowing its nutritional value to be adjusted to the changing needs of the offspring during different seasons.

5. Conclusions

In summary, photoperiod and lambing season significantly influence lactation and metabolic performance in sheep, particularly by modulating hormones such as PRL and insulin and altering the composition of milk components. In contrast, LEP in milk exhibits less seasonal variation, suggesting that its role may be more related to metabolic than photoperiodic regulatory mechanisms. These findings may have practical implications for sheep management, planning production seasons, and optimizing feeding and lighting strategies, which can contribute to improved milk yield and offspring growth.