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Maternal Nesting

By July 7, 2019August 3rd, 2019No Comments


Maternal nesting is the behavior that mother mice carry out when creating a shelter to protect their soon-to-be newborns. In contrast to thermoregulatory or non-maternal nesting, where a mouse is building a nest for non-reproductive purposes, maternal nesting is a specific instinctual behavior that arises strictly within the circumstances of reproduction and birth-giving. 


Maternal nesting is comprised of a complex behavioral chain wherein several behaviors are performed in order to build the nest. A mouse will gather bedding materials and transport them to the nest and then use them to create a compact nest that will later be used to house the newborn pups. Maternal nesting is expected to occur a few days before parturition and lasts for about two weeks while the mouse is lactating and then gradually declines after that.  

Maternal nesting is classified as a nesting behavior since its major outcome is a built nest, but for the specific purposes of reproduction which serves to shelter the newborns, keep them huddled together and warm, and minimize their risk of getting lost. Thermoregulatory or non-maternal nests, on the other hand, are created by the adult mouse for themselves for the purpose of keeping warm and regulating body heat. However, the size of a female mouse’s maternal nest is very likely to correlate with the size of her thermoregulatory nest. 

Maternal nesting is done with the sole purpose of caring for and attending to the newborn pups, It is very important for survival since the quality of the nest is directly related to pup survival. In fact, the pups that are brought up in a nest that is of good condition are more likely to survive in extreme conditions than mice that are brought up in nests of average condition. Furthermore, pup survival is higher in dome-shaped nests as opposed to cup-shaped or flat nests. 

Compared to thermoregulatory nests which are more simple and flat, maternal nests are more complex and bigger. A maternal nest is also sometimes referred to as a “brood nest” in scientific literature. 

Behaviors Comprising Maternal Nesting 

Since maternal nesting is a complex behavior comprised of many other behaviors which make it possible to fulfill the goal of building a nest, many other behaviors are necessary in order to be able to build a nest, including: 

  • Digging. When digging, a mouse is using its fore- and hind-limbs to displace materials. Digging is an essential behavior for nesting and is also often observed when a mouse is burrowing. 
  • Shoveling. Shoveling differs from digging in that the mouse does not use its forelimbs as it does when digging. Instead, when shoveling, the mouse uses its head and snout to move the bedding.  
  • Push-digging. Push-digging is performed in order to move adjacent materials away from the nest instead of towards it. So, a mouse will push-dig any material to the sides of the cage or somewhere away from the nest’s location. Push-digging may be observed in combination with forward movement or locomotion.   
  • Carrying. A mouse will carry materials to the nest location, in order to be able to use those materials to build the nest. This is an important behavior since it is implicated in the actual supply of materials that are necessary for composing a nest. 
  • Fraying. When a  mouse is ripping apart material into smaller pieces by biting or gnawing at the material, it is known as fraying. The resulting torn materials are then used as pieces while nesting and constructing a nest. This resourceful behavior is useful because it demonstrates that a mouse can manipulate materials in order to form shelter and meet one of its basic needs. 
  • Sorting. Sorting is a deliberate action where a mouse is organizing its nesting material. When sorting, the mouse will place the material is specific locations. For example, a mouse may organize bedding material in order based on size, aligning bedding pieces from small to large. 
  • Pulling-in. When pulling-in, a mouse is pulling nesting material toward the nest. The mouse remains in the nest and does not move away from the nest in order to bring in material.
  • Fluffing. Fluffing is a means of enlarging the nest while the mouse is located inside the nest. When a mouse is fluffing, the nest will almost seem like it’s jumping or moving.

Function of Maternal Nesting 

The functions of maternal nesting are to:

  • Provide a safe, nurturing environment. By providing a safe, nurturing environment, pups are able to survive. A safe, nurturing environment offers the pups all the resources they need in order to survive through infancy and grow up into mature mice. 
  • For thermoregulation. Since pups do not have the ability to thermoregulate, they must depend on body heat from their parents and littermates or, insulation from the nest, in order to stay warm. Therefore, nests are a mechanism for warming newborns since they are unable to warm themselves.
  • Position appropriately the pups in order to suckle. A successfully built nest enables the mother to position the pups in such a way that they can ultimately have an easier time suckling. This is an advantage because the nurtured pups are more likely to survive and reach maturity.  
  • To minimize the risk of separation. Since newborn pups cannot see or navigate, the maternal nest is a way to enclose them within a safe environment and minimize their risk of getting separated from the group. This ultimately increases their chance of survival and developing normally. 
  • Support adequate development. Since maternal nests are a means of providing warmth and shelter, they are a crucial component when it comes to ensuring the pup’s development and, ultimately, survival. 

Application of the Behavior 

Since maternal nesting is specific to newborn pups and the reproductive period, it can be observed when an expecting mother is about to deliver newborns. Therefore, the applications of maternal nesting are limited to:

  • After mating. Mice will use more nesting material, in order to build nests, if they have recently mated. Mice are expected to use high amounts of nesting material all the way through parturition, indicating that they are spending more time and focus on nesting. Then, about 10 days after parturition, the amount of nesting materials that they use significantly drops.  
  • When the mouse is pregnant or lactating. A mouse that is pregnant or lactating will demonstrate nesting behavior. Mice that have abnormalities or diseases will nest at lower rates and form less complex nests. More on this in the later sections on Mouse Strains and Abnormalities. 
  • A mouse being in the presence of an infant. If an adult mouse is in the presence of a newborn pup, then its maternal nesting instinct is likely to be triggered. It is possible for mice to begin building nests for newborn pups that are not in their own genetic pool.

Research Techniques for Studying Maternal Nesting

  • Genetic studies. Genetic studies can be used for the purposes of studying maternal nesting in mice by investigating the relationship between genetics and maternal nesting. Through the use of genetic manipulation and genetically modified mice, more can be learned about how genetics influence or determine maternal nesting. Genetic studies ultimately lead to insights regarding how maternal nesting is affected by genetics which may result in scientific breakthroughs and developments on how to improve abnormalities which ultimately influence nesting. 
  • Pharmaceutical studies. Pharmaceutical studies are also a means of studying maternal nesting. By giving mice a supplement of drugs or medicine, researchers are able to identify and study how maternal nesting is affected by certain classes of drugs. Pharmaceutical studies are often combined with genetic studies, which offer genetic disease models, in order to study the impact of drugs specifically on maternal nesting.
  • Behavioral studies. In behavioral studies, researchers subject mice to behavioral tests which are carefully selected based on their validity of studying the desired behavior. Behavioral studies are often combined with elements of genetic and pharmaceutical studies, in order to assess specifically how genetics and drugs affect maternal nesting within a behavioral framework.

Behavioral tests 

Maternal nesting can be assessed throughout a mouse’s reproductive life. Typically, maternal nests will be assessed across many stages. In experimental studies, researchers will use nulligravid, primigravid, and lactating females and compare them to virgin females in order to establish maternal nesting patterns across time. 

3-point Scale and Nest Quality Scores 

Maternal nests can be rated using the following 3-point scale:

  • 0: no nest has been created or a primitive flat nest has been created, the female mouse did not use any of the provided material and a clear nest shape is evident
  • 1: a complex cup/dome-shaped nest has been formed and the female mouse used the nesting material provided
  • 2: a complex hooded nest has been formed meaning that the female mouse used the nesting material provided and the resulting nest is well shaped and its walls form a ceiling. 

The nest quality scores report the qualitative features of the nest, including:

  • Position: where the nest is located in the cage with respect to the cage’s key features (like the cage’s drinking source)
  • Height: the height of the nest with respect to the cage size. Low, if the nest is less than half the cage’s height; intermediate, if the nest is about half of the cage’s height; high, if the nest is more than half the cage’s height. 
  • Additional structures: comments made whether there are other additional structures present, such as small heaps of cotton, other nesting materials, or excrement.
  • Texture: describes whether there is a low, intermediate, or high work done on the nest’s surface. 
  • Shape: classifies the shape of the nest. Cup, if the nest is an open bowl-shaped nest; dome, if there is a hemispheric roof and one opening; tunnel, if the nest has two passageways.

Recording how these features are displayed for the nest provides more information about the nature of the nest. 

Pharmaceutical Studies on Maternal Nesting

Progesterone Increases Nest Weight 

Progesterone is a sex hormone. Administering progesterone subcutaneously through hormone-dipped pellets to mice will lead to a subsequent increase in maternal nest weight and a higher percentage of built maternal nests. In fact, giving progesterone to ovariectomized mice also induces maternal nesting. Also, giving progesterone to non-pregnant mice will lead to the elicitation of maternal behaviors such as maternal nesting.  

Estrogen Dose Suppresses Nest Building

Estrogen, also referred to as oestrogen, is a main female sex hormone. In mice, using the same method as described above, in the ‘Progesterone Increases Nest Weight’ sub-section, through subcutaneously administered pellets, estrogen is associated with suppressed maternal nest building. 

Testosterone Propionate Reduces Maternal Nest Building 

Testosterone propionate is a manufactured medication, designed originally for treating conditions associated with low testosterone levels. In mice, testosterone propionate reduces maternal behaviors, including nesting. Mice that are given testosterone propionate will have lower nest building scores since they create nests that are not as intricate as typical brood nests. 

Lipopolysaccharide Injections Decrease Maternal Nesting

Sickness is associated with altered behavior. Lipopolysaccharides are able to induce behavioral symptoms that are similar to sickness, leading to decreased levels of locomotion, a reduction in social activities, and even anorexia. This is because lipopolysaccharides are a bacterial fragment that can induce sickness by mimicking cytokines’ molecular structure when injected in mice in a controlled setting. Maternal mice that are injected with lipopolysaccharides will demonstrate lower nesting scores than saline-injected mice under a 22°C setting. However, under colder temperatures, maternal mice injected with lipopolysaccharides will build nests that are not significantly different than controls’, implying that sickness modulates behavior under particular circumstances. 

Chlorpyrifos Injections Postpone Nest Building Instinct

Organophosphates (OPs) are an important group of pesticides for which strong human epidemiological evidence has been gathered, that it can cause damage to the nervous system due to acute intoxication. Chlorpyrifos is a particular type of OP pesticide. Chlorpyrifos (CPF) is one of the top-selling insecticides, the most commonly used OP in the USA for the purposes of pest control. Yes, the Environmental Protection Agency has banned its residential use due to potential safety issues of childhood exposure. Newborn pups that have been given injections with CPF affect maternal care. Pups that have been injected with CPF influence maternal care and are associated with postponed nest building. This demonstrates the complexity of maternal nesting by demonstrating that the pups themselves can influence maternal nesting behaviors.

Mouse Strains

Maternal nesting varies across mouse strains. Since mice differ by genetics, the variability in maternal nesting can be assessed and analyzed in such a way that it can be partly explained by genetics. Below are several mouse strains and their expected type of maternal nesting:

Ephrin-A5-/- Mice 

Throughout the central nervous system’s neural circuits, ephrin ligands and Eph families of tyrosine kinase receptors can be found. The Eph families of tyrosine kinase receptors along with the ephrin ligands are involved in regulating animal behavior. Ephrin-A5 is a particular kind of ligand (one of the eight identified ligands for the Eph receptors) which has been found to be expressed in brain regions that are associated with maternal behaviors, such as the preoptic area and the hypothalamus. Ephrin-A5 transcripts can also be detected in the suprachiasmatic nucleus, the arcuate nucleus, and the paraventricular nucleus. Ephrin-A5-/- mice which have the ephrin-A5 genetically knocked out display poorer maternal nesting quality when compared to normal wild-type mice. While pregnant wild-type mice are expected to tear up about 90% of the nesting material and already create a nest by the 6-hour mark of being presented with the nesting material, Ephrin-A5-/- mice leave the nesting material intact, indicating minimal maternal nesting by contrast.   

TPH2-/- mice

Tryptophan hydroxylase-2 (TPH2) is known to catalyze the first and rate-limiting step of serotonin biosynthesis. Mice that have TPH2 mutated (TPH2-/-) show different maternal nesting patterns than TPH2+/+ mice do (mice which still have the gene fully intact). TPH2-/- mothers that have the TPH2 gene mutated have a litter with low survival rates and high incidences of cannibalism. When compared to TPH2+/+ mice, TPH2-/- mice have nests that are poorly constructed, less well-defined and not as tightly gathered. By genetically depleting the brain of serotonin due to TPH2 mutation, subsequent effects on maternal nesting can be noted. 

PET-1-/- Mice

PET-1 is a transcription factor that belongs to the ETS family and is exclusively expressed in brain cells which are a part of the serotonin system. Mice that lack the gene for PET-1 (PET-1-/-) and thus cannot produce this transcription factor show compromised maternal behaviors including poorer nesting skills. One study, by Lerch-Haner et al. established that only 47.6% of PET-1-.- mice will complete their nests compared to 86.5% of PET-1+/+ control mice who complete theirs. Also, only 13.5% of controls will have an incomplete nest while 42.9% of PET-1 deficient mice will have an incomplete nest. Furthermore, almost 10% of PET-1-.- mice will not build a nest at all, an observation that did not occur at all within the controls since they all managed to build at least some portion of a maternal nest.

129Sv Mice

129Sv female mice are quick to start nest building and will begin to nest-build at a sooner point in time than other mouse strains such as C57BL/6J. However, when considering nest building as a part of all of the homecage behaviors that a female mouse exhibits, nest building takes a smaller fraction of this strain’s time, indicating that this behavior does not occur frequently in female 129Sv mice. 

Vitamin D Receptor Knockout Mice

Vitamin D has several key functions within the brain and depends on the vitamin D receptor (VDR) in order to get them done. In humans, there is increasing evidence that links mutations in the VDR with psychiatric conditions. Mice that have had VDR knocked out show many abnormal behaviors, like barbering, and also compromised maternal nest-building abilities. VDR knockout mice are more likely to build a cup-shaped nest (which has a completeness score of about 3 out of 5) while wild-type mice, by comparison, are more likely to create a hooded nest (about a 4 out of 5 of completeness). 



Olfactory bulb removal through surgical means, also known as bulbectomization, results in a condition wherein a pregnant mouse no longer has its olfactory bulb. This is the part of the brain which receives neural inputs regarding odors as sensed by cells located in the nasal cavity. Bulbectomization inhibits nest building in pregnant mice (and also in virgin mice), suggesting that the olfactory bulb is involved indirectly in both thermoregulatory and maternal nesting.  

Fatty-Acid Deficiency

N-3 fatty acid-deficient pregnant mice have lower nest scores than control mice that have an adequate fatty-acid deficient diet. Furthermore, after delivery, about 40% of n-3 fatty acid dams may be expected to attack or neglect their newborns by not nursing them. The deficient dams will have lower levels of brain docosahexaenoic acid (DHA) levels than the dams that follow an adequate diet. 

Vitamin D Receptor Deficiency

A deficiency in vitamin D receptors can influence maternal behavior. Vitamin D, in itself, is a steroid hormone that has multiple important roles in the brain and is mediated by the vitamin D receptors. Without the receptors, maternal nesting becomes compromised and many other abnormalities in maternal behavior occur such as pup neglect and cannibalism. 

Abnormalities of the Serotonergic System 

Serotonin is a hormone and neurotransmitter with various roles in the brain. However, its function on maternal nesting is still not fully understood. Mice that have a mutation in the serotonergic system, such as TPH2-/- mice or PET-1-/- mice, have been shown to have abnormally low scores of maternal nesting which negatively affects pup survival. 

Disease Models of Maternal Nesting

Limited Nesting/Bedding-Material Procedure to Model Early Life Stress 

Limited nesting or the bedding-maternal procedure is a disease model mimicking the early life stress paradigm. A mother is deprived of available nesting materials, thus cannot make a well-formed nest. This, in turn, creates stress on the pups since they do not have a warm, tight nest to grow up in. By modeling stress, researchers can measure how nesting influences upcoming generations of newborns given environmental shortcomings by decreasing the quality of maternal nesting through environmental manipulation. 

Postpartum Depression

Induction through an n-3 fatty acid-deficient diet

Pregnant mice that are fed on an n-3 fatty acid-deficient diet have poorer nest building abilities than pregnant mice following an n-3 fatty acid-adequate diet. Furthermore, about 40% of dams that are on this type of diet are expected to neglect their pups by not nursing them or to attack their newborn pups. Female mice on an adequate diet, have higher levels of brain docosahexaenoic acid than deficient mice. This suggests that such a diet deficient in n-3 fatty acids can be used to induce postpartum disorder in mice and that dietary n-3 fatty acid intake is associated with normal maternal behavior and the prevention of postpartum depression. 

Induction through chronic social defeat

Chronic stress is a known predictor of depressive disorders. Social stress is a particular form of stress that is common in both humans and mice. The majority of mouse models of depression were initially developed for male mice and may not be applicable to postpartum female mice. In order to induce postpartum depression, lactating female dams are exposed daily to a novel male intruder. Subsequently, overall maternal care is reduced which includes nesting and bouts of self-grooming significantly increases in comparison to controls.


  • Nesting behaviors refer to a mouse creating or building a nest for itself or its offspring. 
  • Nesting behaviors refer to any point that a mouse is building its shelter. There are two types of nesting-behaviors known to date: maternal nesting and non-maternal, or thermoregulatory, nesting. 
  • Behaviors that are related to maternal nesting and are important for execution include: digging, shoveling, push-digging, carrying, fraying, sorting, pulling-in, and fluffing.
  • Maternal nesting has many functions and is crucial for: providing a safe and nurturing environment to newborns, providing thermoregulation,  positioning the pups appropriately in order to be able to suckle, minimizing the risk of separation, supporting adequate development. 
  • Maternal nesting can be observed under the following conditions: after mating, when a mouse is pregnant or lactating, or in the presence of an infant. 
  • In order to study maternal nesting, researchers use genetic, pharmaceutical, and behavioral studies, or some combination of them. 
  • Some notable pharmaceutical behaviors have established that:
    • Progesterone increases nest weight.
    • An estrogen dose suppresses ne st building.
    • Testosterone propionate reduces maternal nest building.
    • Lipopolysaccharide injections decrease maternal nesting.
    • Chlorpyrifos injections postpone nest building instinct.
  • Across mouse strains, maternal nesting varies. For example: 
    • Ephrin-A5-/- mice display poorer maternal nesting quality when compared to normal wild-type mice.
    • TPH2-/- mothers that have the TPH2 gene mutated have a litter with low survival rates and high incidences of cannibalism. When compared to TPH2+/+ mice, TPH-/- mice have nests that are poorly constructed, less well-defined and not as tightly gathered.
    • Mice that lack the gene for PET-1 show compromised maternal behaviors including poorer nesting skills.
    • Maternal nest building takes a smaller fraction of 129Sv mouse’s overall cage time, indicating that this behavior does not occur as frequently as expected when the mouse is pregnant. 
    • Mice that have had VDR knocked out show many abnormal behaviors, like barbering, and also compromised maternal nest-building abilities.
  • Abnormalities are known to affect maternal nesting. For example,
    • Bulbectomization inhibits nest building in pregnant mice.
    • N-3 fatty acid-deficient pregnant mice have lower nest scores than control mice that have an adequate fatty-acid deficient diet.
    • Without vitamin D receptors, maternal nesting becomes compromised and many other abnormalities in maternal behavior occur such as pup neglect and cannibalism. 
    • Mice that have a mutation in the serotonergic system, such as TPH2-/- mice of PET-1-/- mice, have been shown to have abnormally low scores of maternal nesting which negatively affects pup survival.
  • Disease models can be used to model abnormalities or diseases of maternal nesting in mice. 
    • Limited nesting/bedding is a disease model mimicking the early life stress paradigm, thereby decreasing the quality of maternal nesting through environmental manipulation. 
    • Postpartum depression is another disease model that affects maternal nesting. It is modeled either through limiting the female mouse to an n-3 fatty acid-deficient diet or exposing her to chronic social defeat. 


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