The Tammar Wallaby and Fur Seal: Models to Examine Local Control of Lactation¹

The lactation cycle of the tammar wallaby. (A) Development of the pouch young from d 6 to 220 of age. (B) The lactation cycle in the tammar has been divided into 4 phases, characterized by changes in milk composition and the sucking pattern of the pouch young. (C) Expression of the major milk protein genes during the lactation cycle. The α-CN, β-CN, α-LA, and β-LG genes are induced at parturition and expression remains elevated for the entire lactation. The genes for ELP (early-lactation protein), WAP (whey acidic protein), and the LLP (late-lactation proteins A and B) are expressed only for specific phases of the lactation cycle.

Concurrent asynchronous lactation. The pouch has been retracted to expose the 4 mammary glands. A 6-d-old young is attached to a teat from a mammary gland secreting phase 2A milk. An older animal at approximately 275 d of age has vacated the pouch and sucks from the elongated teat, which provides phase 3 milk from the enlarged mammary gland. The remaining 2 teats are from quiescent mammary glands.

The “foraging” lactation strategy of the fur seal. The pregnant female arrives at shore to give birth and remains with the pup for approximately 1 wk. For the remainder of lactation, females alternate trips to sea with short trips ashore to suckle their pup.

Northern blot analysis of β-LG, α-CN, and α-LA gene expression in tammar mammary explants. Gene expression is shown in mammary tissue from pregnant tammars at d 24 prior to culture (t₀), explants cultured in media with insulin, cortisol, and prolactin (IFP) for 3 d, explants subsequently cultured in the absence of hormones (NH) for 10 d, and following the reintroduction of IFP for 3 d. The length of culture in days is shown by the subscript. Total RNA (10μg, lower panel) was assayed by Northern blot analysis using [α-³²P]dCTP-labeled cDNA probes for the β-LG, α-CN, and α-LA genes (upper panels). Arrows indicate transcript size in nucleotides and RNA ribosomal bands.

Histological sections of the mammary gland from (A) the lactating Cape fur seal while nursing on shore and (B) the lactating Cape fur seal while foraging at sea. Sections are stained with hematoxylin and eosin. Immature alveoli in the pregnant gland are indicated: lipids (white) and milk proteins (pink) are indicated in the onshore and offshore lactating mammary glands. Magnification ×100. Milk protein gene expression (C): β-CN expression during the Cape fur seal lactation cycle. Analysis of expression using canine Affymetrix chips hybridized to cDNA probes generated from RNA from the pregnant (placental gestation and nonlactating, n=2) and lactating Cape fur seal on shore (n=2) and at sea (n=1; animals in embryonic diapause). (D) Cluster analysis of gene expression profiles from the Cape fur seal mammary gland during different stages of lactation. A total of 1,020 Cape fur seal mammary messenger RNA (mRNA) transcripts were identified with expression levels above an intensity of 250 in any sample type. Hierarchical clustering was conducted using Euclidean distance. Pregnant (placental gestation and nonlactating) and onshore lactating (in embryonic diapause) data represent an average of 2 animals. Offshore data represent a single sample.

(A) Proliferation of wallaby mammary epithelial cells (Wall-MEC) cells cultured in the presence and absence of tammar WAP (tWAP). Results shown are after d 3 of treatment with (+) and without (−) tWAP. Cell growth was analyzed using methods described by Skehan et al. (1990). The vertical bars indicate SEM (n=3, *P<0.05). (B) Expression of the cyclin D1 gene in Wall-MEC cells grown in the presence and absence of tammar WAP (d 3 posttreatment). Expression of the cyclin D1 and GAPDH genes was determined by reverse transcription PCR analyses. The relative expression levels of cyclin D1 and CDK-4 genes in Wall-MEC were quantified by National Institutes of Health Image software by analyzing reverse transcription PCR products. The results show the level of cyclin D1 expression as a proportion of the expression of the housekeeping gene GAPDH.