6In Vitro Lactogenesis Control contains insulin and T3. Note minimum requirements are Insulin (or IGF-I), prolactin and cortisol. Prolactin alone causes only a minimal response. Cortisol has no effect. However, the hormones show a substantial synergy.The concentrations of prolactin used are similar to what is found in blood at calving.Control contains insulin and T3.
7Progesterone on Lactogenesis Progesterone inhibits lactogenesis. BUT: has less effect once lactation established. Induction of lactation decreases progesterone receptors in the mammary gland. The levels that inhibit lactogenesis are similar to those in pregnant cows.Control contains insulin,cortisol and T3.
8GH on LactogenesisGH has very little effect on lactogenesis. Major effect on lactation appears mostly indirect, on body metabolism. Growth hormone in cattle is normally about 10 ng/ml. The 100 ng/ml concentration would be seen sometimes, but th higher concentration would never be seen.
9Hormone Priming on Lactogenesis Bovine mammary tissue was transplanted to athymic “nude” mice as described for placental lactogen. Mice were treated with control or estrogen + progesterone (E+P). They were then injected with vehicle and hydrocortisone + prolactin (F+Prl). Note that F+Prl induced milk protein synthesis to a greater extent if tissue was primed with E+P.From:Sheffield, l.G. and C.W. Welsch, J. Dairy Sci, 71:75-83, 1988.
10Extracellular Matrix on Lactogenesis HC11 cells (a mouse mammary cell line) cultured to confluency (each cell contacting a neighbor) with or without epidermal growth factor (EGF) or on a laminin rich matrix and lactation induced with insulin, hydrocortisone and prolactin. Without EGF, little milk protein was present. With EGF, considerable milk protein was induced by prolactin.EGF is very similar in its effects as TGFa. In fact, they bind the same receptor.EGF induce the expression of the extracellular matrix protein laminin. Replacing EGF pre-treatment with laminin coating on the cell culture dishes gave the same effect as adding EGF. Importantly, the laminin content of the matrix is increased during pregnancy, probably in preparation for subsequent lactogenesis.
11Model of LactogenesisDuring pregnancy, hormones and growth factors induce growth, alveolar development and a laminin rich extracellular matrix, in preparation for lactation. Progesterone inhibits induction of lactation, so milk protein synthesis is low until it is needed.At calving, progesterone drops, cortisol and prolactin increase and these, together with the laminin rich matrix, induce milk protein synthesis. Also, many other genes needed for milk synthesis are also increased.Also, at lactogenesis, progesterone receptors decline to very low levels. This allows the lactating cow to be pregnant without a dramatic decline in milk production. There is some decline relative to non-pregnant cows, but not nearly as much as if the gland was as responsive to progesterone as it is in heifers.
19Insulin, Growth Hormone and Cortisol Through Lactation
20OxytocinOxytocin is a 9 amino acid long peptide. The amino acid structure of oxytocin is:Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Leu-GlyIt has a molecular mass of 1007 daltons. Oxytocin has a disulfide bond between the two cysteines. Reduction of the disulfide bond inactivates oxytocin. One IU (international Unit) is approximately 2 micrograms of pure peptide.
21Oxytocin SynthesisOxytocin is synthized in the hypothalamus in specific nuclei, the paraventricular nucleus and the supraoptic nucleus in the hypothalamus. [A cluster of nerve cells in the brain is often called a nucleus. This is different from the nucleus of a single cell.] Neurons in these hypothalamic nuclei synthesize the oxytocin precursor and package it into vesicles. Oxytocin is initially synthesized as a large molecular weight precursor which also consists of the oxytocin-carrier peptide neurophysin. The precursor is proteolytically cleaved in the neuron in the oxytocin-containing vesicle to yield oxytocin bound to neurophysin. The oxytocin-neurophysin complex is the intracellular storage form of oxytocin.The oxytocin-containing vesicles are transported from the cell body (which is in the hypothalamus), down the axons to the neuron endings in the posterior pituitary. This is called the hypothalamo-neurohypophysial tract. The oxytocin-neurophysin complex is stored in neurosecretory granules called herring bodies in the axon ending.
22Pituitary GlandNote that the posterior pituitary gland is actually part of the brain. Nerves originate in the hypothalamus and have endings in the posterior pituitary.Oxytocin is synthesized in the hypothalamus, transported down the axons and stored in the nerve endings in the posterior pituitary. Nerve signals associated with miling cause its release, and it is then taken up by the blood for transport to the udder.The posterior pituitary is also called the Neurohypophysis, to indicate it is nerve tissue.
23Note that oxytocin is release rapidly, and has a very short lifespan in blood. Its half-life is about 1 minute.This is why the milking machine should be attached shortly after udder stimulation (about 1 minute).The 1 minute wait allows time for oxytocin to circulate from the pituitary to the udder.
25Alveolus Stained to Show Myoepithelium MyoepithelialCellThis is a photograph of an alveolus stained to show the myoepithelial cells in detail. Most staining methods do not show them as well. Note that they do not completely surround the alveolus. The basement membrane was digested away to show this. They lie on the same side of the bsement membrane as the epithelial cells.This is a major error in some text (mostly more general physiology texts and some medical works). They mistakenly place the myoepithelium outside the basement membrane. This comes from the notion that the prefix myo, which refers to muscle, indicates that they are of mesodermal origin. In reality, they are specialized EPITHELIAL cells (not the last part of the word). They arise from ectoderm, not mesoderm. They differentiate from the other epithelial cells as the gland develops.
26Milk Letdown ReflexThis is a review for most who have had introductory dairy science.Stimulation of the udder causes a nerve signal to travel from the udder, througn the inguinal nerve, which is the major nerve supplying the udder. It then travels up the spinal cord to the brain. In the brain, it stimulates the hypothalamus. The hypothalamus contains nerve cell bodies which produce oxytocin. Oxytocin travels down the axones, which end in the posterior pituitary, where oxytocin is stored in the nerve endings. When stimulated, these nerves release oxytocin into the blood. The venous blood draining the posterior pituitary is carried to the heart, then lungs, back to the heart and through the aorta. When it reaches the udder, oxytocin binds its receptor in the myoepithelial cells, causing them to contract. This raises intramammary pressure, causing milk ejection.The next several slides will cover each of these processes in more detail. This will include material on the nervous system, relevant material on the endocrine system and end with material on milking machines and how they operate.
27Milk Letdown Alveolar Contraction Oxytocin stimulates CONTRACTION of the myoepithelial cells.This increases intramammary pressure and aids in milk removal. Necessary because small vessels, like mammary ducts, have high resistance to fluid flow (Pascall’s law from physics).
28Oxytocin Release and Half-Life It is estimated that the bovine pituitary has about 800 micrograms of oxytocin. This is about 40X what is in the blood under resting conditions. Only about 1/3 of pituitary oxytocin is released at a milking.Oxytocin receptors on myoepithelial cells can respond to very low levels of oxytocin.Oxytocin has a short half-life in the blood = 0.55 to 3.6 min. This means that the removal of milk by machine or by nursing must be closely timed with stimulation of the teats.
29Factors Modifying Milk Letdown Autonomic nervous systemStress gives epinephrine releaseInhibits oxytocin releaseInhibits myoepithelial cell contractionInhibits blood flow to udderConditioned reflexLetdown in response to sights, sounds associated with milkingThis is why the milking environment should be non-stressful.
31Adrenal Medulla Posterior Pituitary Udder Vasculature Ihnibits oxytocin releaseInhibitsBloodFlowEpinephrineMyoepitheliumThe adrenal medulla is a major (but not only) mediator of acute stress. It is composed of nerve tissue, and is part of the sympathtic nervous system. When sympathetic stimulation reaches the adrenal medulla, it releases epinephrine into circulation. This acts as a hormone, affecting a variety of tissues. It has the same effect as norpinphrine, the neurotransmitter of the sympathetic nervous system.Epinephrine decreases blood flow to the udder, by stimulating contraction of vascular smooth muscle in the udder. Note that in some tissues, like bronchial tissue in the lung, it causes relaxation. As discussed before this is because different tissues have different types of receptors and signaling pathways.Epinephrine also blocks oxtocin release from the posterior pituitary, and inhibits oxytocin action on the myoepithelium.Furthermore, the udder is innervated by sympathetic fibers. Most of these terminate at or near smooth muscle in the blood vessels. When their activity increases, the vascular smooth muscle of the udder contracts and blood flow is decreased.InhibitsContractionMedullaAdrenal
32Milking ApparatusThe teat cup consists of a stainless steel or hard plastic shell, and a flexible liner.These attach to the claw, a small receiving chamber for the milk from each quarter. The vacuum line branches into the 4 vacuum lines supplying each teat cup. The milk lines from the teat cups are attached to the claw, which is attached to the milk hose, which takes the milk to the milk line that leaves the parlor.The milk lines are under constant vacuum. The smaller vacuum lines provide pulsation, as described in the next slides.The chain is for automatic takeoff of the machine when milk flow rate fals below a preset level. This prevents exposing the teat to vacuum after milking is essentially over (termed overmilking).
33Teat Cup StructureThe teat end is under constant vacuum, which removes milk. The space between the shel and liner is called the pulsation chamber. It is under vacuum intermittently. When it is under vacuum, milk flows, but milk flow stops when air is admitted. This is termed pulsation. This is important for teat health, as exposure to constant vacuum can cause tissue damage.Be sure to know that vacuum milks cows, not pulsation.BUT:Pulsation is necessary for proper teat healthAnd may also contribute to oxytocin release or a secondary oxytocin releaseSo milking efficiency is improved with pulsation.
39Removing Residual Milk Oxytocin injectionsExpensiveNot approved useMachine strippingHigh incidence of liner slipsIncreases mastitis riskUdder massageSecond oxytocin release.Udder massage (even briefly) is the best here. Oxytocin is expensive and not approved for this use. Machine striping is to dangerous.Remember: this milk is higher in fat, which results in increased milk price. So, brief udder massage can not only give a bit more milk, that milk is more valuable than the earlier milk.