Name: hand-rearing method for infant marmosets
Uploaded: 2023-06-09T13:40:00.000+00:00
Duration: 4 min 52 s
Description: Watch this Scientific Journal Video about Marmoset Research - Scope and Challenges at JoVE.com

The authors would like to thank Mingxuan Li for his editing of the grammar and polishing of the early version of this manuscript. This work was supported by the Zhejiang Province Natural Science Foundation of China (LD22H090003); the Natural Science Foundation of China (32170991 and 32071097), STI2030-Major Projects 2021ZD0204100 (2021ZD0204101) and 2022ZD0205000 (2022ZD0205003); and the MOE Frontier Science Center for Brain Science &#38; Brain-Machine Integration, Zhejiang University.

All the experimental procedures were approved by the Animal Use and Care Committee of Zhejiang University and followed the National Institutes of Health (NIH) guidelines.
1. Housing and husbandry44
<ol>
	<li>Set the colony room with a 12 h:12 h day/night cycle, the temperature at 26-28 &#176;C, and the relative humidity at 45%-55%.</li>
	<li>Pair male and female marmosets at 2-6 years old, and hold them in cages (850 mm x 800 mm x 800 mm) with sufficient space and fresh air with a 24 h ventilation system.</li>
	<li>Provide resting boards, swings, perches, and hammocks in the cages.</li>
	<li>Feed the pair of marmosets with fresh water and 30-40 g of food twice a day, including cereal, eggs, sweet potatoes, honey, fruit, vegetables, and mealworms. 
	&#8203;NOTE: Veterinarians and experimenters must inspect the animal facility at least once per day to ensure that any sick individuals are diagnosed and treated immediately.</li>
</ol>
2. Preparation before the birth of the marmoset infants
<ol>
	<li>Care for the pregnant marmosets 
	NOTE: The time of conception was diagnosed by palpation (typically 10-20 days after the start of the embryonic period), and we also referred to the reproductive history of the animal subjects.
	<ol>
		<li>Provide a larger space and minimal human disturbance for the breeding pairs.</li>
		<li>Feed the breeding pairs with additional food such as mealworms, eggs, yogurt, and dried fruit to guarantee nutrition for the females.</li>
		<li>Take care of and frequently check the pregnant marmosets to prepare for their parturition. 
		NOTE: The gestation period of the marmoset is estimated to be 148 days &#177; 4.3 days45.</li>
	</ol>
	</li>
	<li>Prepare the following items: an animal incubator (855 mm [W] x 470 mm [L] x 440 mm [H]), disposable diaper pads (M/L sizes), baby wipes, plush toys (Figure 1A), toy rollers (Figure 1B), climbing frames (Figure 1B), blankets (10 cm &#1093; 10 cm, Figure 1C), and an electronic scale (precision of 0.2 g, Figure 1F). 
	&#8203;NOTE: To avoid animal hanging, the plush toys should not have loop structures.</li>
	<li>Food and feeding appliance preparation
	<ol>
		<li>Prepare the following items: baby formula (suitable for 0-12 months of age), baby rice paste (suitable for 0-6 months of age), electric water kettle, beaker (100 mL), heating pad, plastic weighing dish (80 mm x 80 mm x 22 mm, Figure 1D), sterile centrifuge tubes (50 mL), disposable sterile syringes (1-5 mL), intravenous injectors (for custom-made feeding nipples) (Figure 1E), and swabs (80-100 cm). 
		NOTE: To make a nipple for feeding, a cut is made 1 cm from the end of an intravenous injector attached to a syringe (Figure 1E).</li>
	</ol>
	</li>
	<li>Recording form preparation
	<ol>
		<li>Prepare a form, usually several pages long, for each infant marmoset to record basic information such as the name, birth date, birth weight, parents, other basic information of interest such as the head circumference and tail length, breeding information such as the breeding date and time, the amount (mL) of the food intake, the defecation state (yes/no, hard/loose), and the incubator temperature and humidity. 
		&#8203;NOTE: Usually, the body weight is measured and recorded twice a day, once before the first meal and once before the last meal.</li>
	</ol>
	</li>
</ol>
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/65296/65296fig01.jpg" /> 
Figure 1: Photos of the items in the incubator and the feeding tools and accessories. (A) Plush toys; (B) toy roller and climbing frames; (C) blanket; (D) plastic weighing dish; (E) intravenous injector and syringe with a custom-made feeding nipple; (F) electronic scale; (G) caregiver with personal protective equipment. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/65296/65296fig01large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
3. Hand-rearing procedure
<ol>
	<li>Clean and sterilize the room ahead of the due date.
	<ol>
		<li>Spray hypochlorous acid or 75% ethyl alcohol on the floor and table, leave for 30 s, and then wipe the table, and mop the floor.</li>
	</ol>
	</li>
	<li>Set the temperature of the incubator at 35 &#176;C and the humidity at 40%. Usually, to simulate the basic temperature requirement of infant marmosets, as Table 1 shows, before postnatal day 14 (P14), keep the incubator temperature at 35 &#176;C, and from P15 on, lower the temperature by 0.5 &#176;C every 3 days. Keep the humidity at 40%-45% inside the incubator, which is close to the colony humidity and keeps the fur of the infants dry.</li>
	<li>Tile a disposable diaper pad to cover the chassis of the incubator.</li>
	<li>To minimize the stress of the infants, put a couple of blankets and plush toys in the incubator ahead of introducing the infant marmosets, which tend to mimic the adult marmosets. 
	NOTE: The blankets and plush toys are put in the home cage of the breeding pairs for 1 day before use.</li>
	<li>Put the infant marmosets into the incubator and place them on the plush toys once they are separated from their parents in the home cage. 
	NOTE: To avoid social isolation and in accord with animal welfare, generally, two infants are chosen together for hand-rearing.
	<ol>
		<li>Wear sterilized personal protective equipment (PPE, Figure 1G) before feeding.</li>
		<li>Warm a couple of blankets to 35 &#176;C.</li>
		<li>Gently hold the infant marmoset with warm blankets, and obtain the weight of the animal as the birth weight.</li>
		<li>Transfer the infant marmoset into the incubator with warm blankets.</li>
		<li>Take records, as mentioned in step 2.4.</li>
	</ol>
	</li>
	<li>Blend food ingredients and feed the infant marmoset.
	<ol>
		<li>Dissolve 5 g of baby formula in 30 mL of 50 &#176;C boiled water in a 50 mL sterile centrifuge tube. 
		NOTE: Infant marmosets at different postnatal ages need different food recipes. Table 2 includes the different dosages of baby formula, rice paste, and water at postnatal ages from P1 to P60. The dosage is usually enough for 1 day. Blend the ingredients before the first meal, and store the rest of the food in a 4 &#176;C fridge. Heat the food to 30-35 &#176;C for each meal.</li>
		<li>Take 1 mL of food with a 1 mL syringe, and lid the syringe with a custom-made feeding nipple. 
		NOTE: Select a syringe of the proper size, referring to Table 2.</li>
		<li>Keep the food temperature at 30-35 &#176;C with the heating pad.</li>
		<li>Warm the hands before feeding, and gently hold the infant marmoset with a warm blanket in one hand in the incubator.</li>
		<li>Put the feeding nipple into the mouth of the infant marmoset while the head of the infant marmoset is softly held with the thumb and index finger of the holding hand of the caregiver, and slowly push the food out of the syringe at a constant velocity. 
		NOTE: Never push the food faster than the infant marmoset swallows. Pushing fast can cause choking, with food coming out from the nose through the throat. This can cause diseases such as pneumonia, which can even lead to death. If the food overflows, the infant marmoset will struggle. Whenever this occurs, stop feeding and wipe the food off the animal&#39;s face carefully. Continue to feed after the marmoset starts behaving normally.</li>
	</ol>
	</li>
	<li>After the infant marmoset consumes a proper amount of food, wipe its anus with a swab with warm water, which both cleans the anus and promotes defecation.</li>
	<li>Observe the animal for a few minutes, and check the locomotion and defecation of the animal.</li>
	<li>Record the feeding time, the amount (mL) of the food intake, the defecation state (yes/no, hard/loose), and the incubator temperature and humidity. 
	NOTE: Wrap the infant marmoset with warm blankets during weighing to avoid the infant getting cold or injured.</li>
	<li>Keep the chassis of the incubator clean by picking up solid feces or changing to a new disposable diaper pad.</li>
	<li>Before P50, feed the infant marmoset following steps 3.3-3.7, and use the dosage of food ingredients and the feeding time and frequency shown in Table 2.</li>
	<li>From P50 on, the infant marmoset is usually ready for voluntary eating.
	<ol>
		<li>Use plastic weighing dishes instead of syringes. Prepare the food by mixing food ingredients directly in the dish; refer to Table 2 for the amounts.</li>
		<li>Put the food dish in the incubator, and fix the bottom in case it is turned over. Observe for a few minutes to make sure the infant marmosets eat the food. For the first few times, guide the animal to eat voluntarily by luring the animal to the food dish and guiding its mouth to touch the food several times. 
		&#8203;NOTE: Never let the animal&#39;s nose touch the food. Usually, the animal learns to voluntarily eat in 1 day.</li>
	</ol>
	</li>
</ol>
4. Acclimation before the return of the infant marmosets to the colony
NOTE: Usually, the hand-rearing is finished when the infant marmosets learn to eat by themselves. There are a few adaptation procedures to be carried out before they are returned to the home cage in the marmoset colony.
<ol>
	<li>Move the infant marmosets from the animal incubator to small cages (45&#160;cm x 45&#160;cm x 40 cm), which are similar to a birdcage. Hang a water bottle (50 mL) on each small cage.</li>
	<li>Transfer the small cages with the infant marmosets to the colony, and position them close to the family cage.</li>
	<li>Feed the infant marmosets separately with a plastic weighing dish for 1 week, mixing the food according to the daily recipes prepared for the whole colony.</li>
	<li>Make a record of the body weight and defecation status once per day.</li>
</ol>
5. Infant marmosets returning to the family cage
NOTE: After living in the small cage for 7-10 days, the infant marmosets usually adapt to the colony environment well and exhibit no more anxiety.
<ol>
	<li>Put the infant marmosets back into the family cage in the morning.</li>
	<li>Observe the animals for at least 15 min to make sure there is no biting, fighting, or chasing between the family members and the new entrants. 
	NOTE: If biting, fighting, or chasing occurs, separate the infant from the others as soon as possible; and try to return the infant to its family one more time on another day. If the failure occurs again, select another family to foster it. The family groups that have rich parenting experience are the first choice for fostering. Put a plush toy in a new cage to accompany the infant monkey if no family accepts it.</li>
	<li>Stop feeding the infant marmosets separately, and start feeding them using the diet for the colony.</li>
	<li>Pay close attention to the activity of the infant marmosets for 1 week after they return to the family cage.</li>
	<li>Measure the body weights of the infant marmosets every 2 days, and make a record. If they lose weight, feed them extra nutritional food by syringe in the family cage.</li>
	<li>Provide daily care for the infant marmosets, as for the marmosets in the colony.</li>
</ol>

Enhancing Survival and Development of Marmosets Using a Hand-Rearing Approach

The authors have no conflicts of interest to disclose.

The common marmoset is a very useful NHP model for biomedical and neuroscience research. However, marmoset resources are too limited to meet the rapidly growing needs. In this work, we have developed a hand-rearing method that not only increases the survival rate of marmoset infants but also provides an opportunity to study their postnatal development. This hand-rearing method is practical and easy to learn and is, therefore, easily applicable to other labs working with common marmosets.
Some congenital defects usually develop in the first 2 weeks of life. Until now, we have identified several cases of newborn marmosets with innate defects on the eyelid, the lung, the intestine, and the brain, and they rarely survive until P14. Therefore, P1-14 is a critical period for screening out healthy newborn marmosets that can be raised by the hand-rearing method. In addition, if infant marmosets are dropped from the body of their parents due to negligence or lack of milk, they cannot survive. Thus, the hand-rearing method helps to save their lives. In this work, without hand-rearing, the survival rate was 45% (n = 13/29), and it increased to 86% (n = 25/29) after we applied the hand-rearing method.
There are several critical points to note. The caregivers must be patient during feeding; pushing the syringe too fast is not good, as it does not allow the infant marmosets to swallow, which may lead to choking or ileus and, thus, impair the respiratory system or the digestive system of the animals. The caregivers must be gentle and quiet during feeding to minimize the stress to the infant animals, because stress could significantly affect the gut microbiome and is associated with chronic diarrhea in marmosets47,48. During the hand-rearing, all the people near the infant marmosets have to behave gently throughout. Sometimes, opening the door of the incubator in a hurry makes sudden loud sounds, which can frighten the infants and lead to a stress reaction.
The body weights of the animals must be monitored every day, because body weight is a key indicator of an animal&#39;s health status. Underfeeding, overfeeding, malnutrition, or dyspepsia can be detected quickly by a change in body weight. It has been previously reported that marmosets with low body weights are more prone to concurrent bone and gastrointestinal diseases49. Generally, the body weights of the infant marmosets remained constant in the P1-5 stage and gradually increased by 1-4 g per day when using the hand-rearing method. From P5 onward, if no increase or a gradual loss of body weight over 2 days is observed, the caregivers must be alert and check the food ingredients, as well as the body temperature and excretion status of the animals. It is useful to add probiotics in this case.
It is not necessary to feed the animals overnight. We tried to feed the infant marmosets every 2 h to 4 h around the clock and found that it did not promote the growth of the infant marmosets (data not shown). Lacking adequate physical exercise or overfeeding could induce metabolic syndromes, such as obesity, which has been linked to dyslipidemia, altered glucose metabolism, and insulin resistance in marmosets50,51,52. The infants must be kept warm during feeding and weighing, especially in the first few days after birth. We found that one infant had diarrhea after it caught a cold; diarrhea rapidly weakens the animal&#39;s vitality and reduces the body weight.
The grip strength and body temperature are the main indicators of the health status of infant marmosets, and these must be monitored carefully. Whenever an infant marmoset is being fed, it grips the blanket or the feeder&#39;s finger. The grip strength is usually weakened when the animal has diarrhea. Once this happens, one should add pea-sized probiotics to the food and feed the affected infant for 3 days until the diarrhea is gone. If the diarrhea lasts over 3 days, one should add montmorillonite powder instead of probiotics. In contrast, if the infant is constipated, one should gently wipe its anus with a swab dipped in warm water to promote defecation, and lactulose should be used if necessary. In addition, the body temperature of the infant marmoset is usually higher than that of humans. If the animal&#39;s temperature is lower than normal, which is often associated with a weakened grip strength, the animal&#39;s behavior must be monitored closely and the feeding adjusted accordingly.
When the infant marmosets start to climb, they must be provided with climbing frames and toy rollers in the incubator under the supervision of the breeder. The animals are allowed to practice climbing before and after each meal from P25 onward. At P50, the infant marmosets can climb and jump well, and this is a prerequisite for voluntary eating. Infant marmosets like to grind their teeth by biting blankets, plush toys, or disposable diaper pads, but these may form small loops that threaten their lives due to self-strangulation. Hence, torn pieces and thread should be cleaned promptly to avoid injuries or deaths.
Sometimes, the feeding procedure can be modified slightly due to differences in appetite, vitality, and environmental conditions. If an infant marmoset is strong enough, it will probably start to practice climbing before P25, learn voluntary eating before P50, and even go back to the family before P60. Meanwhile, we lower the temperature by 0.5 &#176;C in the incubator every day or every 2 days so that the infant marmoset can adjust to the same temperature as that in the colony before it eventually goes back to the cage. If an infant marmoset has a poor appetite for a certain period, less food must be provided per meal but at a higher frequency per day, and the incubator must be kept warmer than the colony for a longer time. In the current study, 88.9% (n = 16/18) of infants were successfully accepted by the birth parents, 11.1% (n = 2/18) were fostered by other families, and none were rejected.
Studying juvenile marmosets with different family structures reveals differences in their social and mental development; indeed, social interactions between parents and children and between peers during development potentially sculpt social brain function53,54,55. Therefore, whether one or two animals are selected for hand-rearing depends on the aims of the experiment. Our lab usually hand-rears two infants, if possible, to both meet animal welfare demands and help more infants survive. The collaborative rearing method for marmoset triplets43 is also a good choice. Hand-reared marmosets with limited parental interactions may not be suitable for experiments for the study of social behavior.
In conclusion, the hand-rearing method in the current study provides instructions on how to feed infant marmosets from P1 to P60. With this protocol, it takes ~15 min for an experienced breeder to manually feed one infant marmoset. The successful operation depends on multiple factors, including the proper temperature of the food and the environment, the feeding time and technique, and the excretion of the animals after meals. This method is feasible and easy to learn. It has been continually optimized with 5 years of practice. This method may be applied to other laboratories working with common marmosets.

The common marmoset (Callithrix jacchus) is a small and arboreal New World monkey originating from South and Central America. The use of marmosets in biomedical research has grown rapidly over the past decades due to several key advantages of marmosets compared with other non-human primates (NHPs), including their smaller body size, easier handling and breeding in captivity, shorter gestation time, earlier sexual maturation, and lower zoonotic risks1,2,3,4,5,6. The common marmoset has a similar brain structure and brain function to humans and displays a rich repertoire of vocalizations and highly social behavior with rich emotions. It is a compelling NHP model for different types of neuroscience studies, such as studies on sensory processing7,8,9,10,11,12,13,14, vocal communication15,16,17,18,19, models of spinal cord injury20,21,22,23, Parkinson&#39;s disease24,25,26,27,28, and age-related diseases29. Compared with other NHPs, the common marmoset has a relatively high reproduction rate, which is potentially useful for transgenic modification30,31,32. This primate is also widely used in pharmacology, angiography, and pathogen and immune studies33,34,35,36,37,38,39. However, the supply of marmosets remains very limited, especially in China, and cannot meet the rapidly growing needs of scientific research.
In marmoset colonies, the adult animals are fed once or twice per day, and a few institutions alter the diet for juvenile marmosets40. Generally, infant marmosets usually grasp firmly onto the body of the father or elder siblings for daily care and are handed to the mother several times per day for milk. Some female marmosets give birth to triplets, and in this case, one or two infants cannot survive due to a lack of milk; moreover, some parents do not take care of their infants because they lack nursing experience or for other unknown reasons. This is a big loss for many laboratories. A few studies have reported methods of nutrition management for adult marmosets in captive settings40,41,42 utilizing foods and formulas with different macronutrient compositions, vitamins, and minerals, as well as different feeding protocols for enrichment (mashed, gelled, purified, or canned)2,41. One previous study reported a collaborative rearing method for marmoset triplets43, in which caregivers take one infant per day, hand-feed it throughout the day, and exchange it for another of the triplets on the next day. Although this method allows the infants to have parental care, it requires an experienced caregiver to grab the infant from the body of the parents every day and is labor-intensive. So far, no study has reported a detailed, step-by-step hand-rearing method for newborn marmosets.
The goal of the current study is to provide a hand-rearing method for those interested in marmoset development but with limited resources. In contrast to the previous collaborative rearing method43, the current method is an alternative that causes less disturbance to the infant&#39;s family and is easy to learn. Based on the basic rules of breastfeeding and 5 years of practice, this paper describes a hand-rearing method for raising infant marmosets that includes the preparation of the food, a timetable for feeding, the configuration of the temperature and humidity of the animal incubator, as well as the adaptation of the infant animals to the colony environment.

<table><tbody><tr><td>animal incubator</td><td>RCOM, Korea</td><td></td><td>MX - BL600N, 855 mm (W) x 470 mm (L) x 440 mm (H)</td></tr><tr><td>baby milk powder</td><td>Meadjohnson, America</td><td></td><td>suitable for 0-12 months of age, executive standard - GB25596</td></tr><tr><td>baby rice paste</td><td>HEINZ, China</td><td></td><td>suitable for 0-6 months of age, executive standard - GB10769</td></tr><tr><td>baby wipes</td><td>babycare, China</td><td></td><td>soft</td></tr><tr><td>beaker</td><td>ShuNiu, China</td><td></td><td>100 mL</td></tr><tr><td>blankets</td><td>Grace, China</td><td></td><td>10 cm &amp;times; 10 cm, soft</td></tr><tr><td>climbing frame</td><td>WowWee, China</td><td></td><td>firm and no small circular structures</td></tr><tr><td>disposable diaper pads</td><td>Hi Health Pet, China</td><td></td><td>either M or L size</td></tr><tr><td>disposable sterile syringe</td><td>Cofoe, China</td><td></td><td>1 mL, 2.5 mL, 3 mL, 5 mL, 10 mL</td></tr><tr><td>electronic scale</td><td>YouSheng, China</td><td></td><td>measuring range from 0 to 6,000 g with precision of 0.2 g</td></tr><tr><td>intravenous injector</td><td>HD, China</td><td></td><td>0.55 mm x 20 mm needle</td></tr><tr><td>kettle</td><td>FGA, China</td><td></td><td>warm-keeping kettle 1,500 mL</td></tr><tr><td>lactulose</td><td>BELCOL, China</td><td></td><td>to solve constipation</td></tr><tr><td>plastic weighing dish</td><td>SKSLAB, China</td><td></td><td>80 mm x 80 mm x 22 mm, used as a bowl</td></tr><tr><td>plush toy</td><td>Lebiyou, China</td><td></td><td>soft</td></tr><tr><td>probiotic powder</td><td>G-Pet, China</td><td></td><td>to regulate gastrointestinal environment</td></tr><tr><td>sterile centrifuge tube</td><td>NEST, China</td><td></td><td>50 mL</td></tr><tr><td>swab</td><td>OYEAH, China</td><td></td><td>80 - 100 mm</td></tr><tr><td>toy roller</td><td>WowWee, China</td><td></td><td>firm and no small circular structures</td></tr></tbody></table>

hand-rearing method for infant marmosets

The common marmoset (Callithrix jacchus) is a small and highly social New World monkey with high reproduction rates, which has been proven to be a compelling non-human primate model for biomedical and neuroscience research. Some females give birth to triplets; however, the parents cannot raise all of them. To save these infants, we have developed a hand-rearing method for raising newborn marmosets. In this protocol, we describe the formula of the food, the time for feeding, the configuration of the temperature and humidity, as well as the adaptation of the hand-reared infants to the colony environment. This hand-rearing method significantly increases the survival rate of marmoset infants (without hand-rearing: 45%; with hand-rearing: 86%) and provides the opportunity to study the development of marmoset infants with similar genetic backgrounds raised in different postnatal environments. As the method is practical and easy to use, we anticipate that it could also be applied to other labs working with common marmosets.

Body weight is a key index of animal body development and is used as an indicator of the health status of the marmosets in this protocol. In this work, the body weights of the hand-reared animals increased gradually with age (Figure 2A, n = 16), similar to the weights of newborn infants in a previous study46. To minimize the disturbance to the breeding families in the colony, we did not weigh the infant marmosets in the colony every day. We obtained the weights of the parent-raised animals 1 month after birth and later, and these were compared with the weights of the hand-reared infants at the same ages. There were no significant differences in the body weights between the parent-raised infants and the hand-reared infants (P30, hand-rearing group, 52.25 g &#177; 2.10 g, n = 18; colony group, 57.34 g &#177; 2.77 g, n = 7; t = &#8722;1.3417, P = 0.1928; P60, hand-rearing group, 91.76 g &#177; 3.44 g, colony group, 93.06 g &#177; 4.68 g; t= &#8722;0.2019, P = 0.8424; Student&#39;s t-test) (Figure 2B).
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/65296/65296fig02.jpg" /> 
Figure 2: Body weights of the hand-reared infant marmosets and the parent-raised ones in the colony. (A) The body weight of each hand-reared infant. The gray lines indicate individual body weights, and the orange curve indicates the population average. (B) Comparison of the body weights between the hand-reared infants (orange) and the parent-raised infants (green) at P30 and P60. The gray circles represent individuals. Data shown as Mean &#177; SEM. ns indicates no significant difference. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/65296/65296fig02large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
Table 1. Configuration of the temperature in the animal incubator. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/65296/65296_Table1rev-20230220RV4.xlsx" target="_blank">Please click here to download this Table.</a>
Table 2: Feeding recipes. The columns under &#34;food ingredients&#34; indicate the milk recipe for 1 day. For example, at postnatal days 1-14, we mix 5 g of baby milk powder and 30 mL of water before the first meal and take 0.5-1.5 mL of this milk each time for feeding. The dosages of &#34;baby milk powder&#34; and &#34;rice paste&#34; are for the powder forms. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/65296/JoVE-Table2-FeedingRecipes-20230502R3.xlsx" target="_blank">Please click here to download this Table.</a>

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Hand-Rearing Method for Infant Marmosets

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Pharmacogenetic research benefits first-hand from the abundance of information provided by the completion of the Human Genome Project. With such a ...

Method for Whole Mount Antibody Staining in Chick

The chick embryo is a valuable tool in the study of early embryonic development. Its transparency, accessibility and ease of manipulation, make it an ideal tool for studying antibody expression in developing brain, neural tube and somite. This video demonstrates the different steps in whole-mount antibody staining using HRP conjugated secondary antibodies; First, the embryo is dissected from the egg and fixed in paraformaldehyde. Second, endogenous peroxidase is inactivated; The embryo is then exposed to primary antibody. After several washes, the embryo is incubated with secondary antibody conjugated to HRP. Peroxidase activity is revealed using reaction with diaminobenzidine substrate. Finally, the embryo is fixed and processed for photography and sectioning. The advantage of this method over the use of fluorescent antibodies is that embryos can be processed for wax sectioning, thus enabling the study of antigen sites in cross section. This method was originally introduced by Jane Dodd and Tom Jessell 1.

This video demonstrates whole mount immunohistochemistry, a method by which the spatial and temporal expression pattern of an antigen can be visualized in young chick embryos. This method was originally introduced by Jane Dodd and Tom Jessell.

The chick embryo is a valuable tool in the study of early embryonic development. Its transparency, accessibility and ease of manipulation, make it an ...

A Common Marmoset Model of Mother-Infant Intervention for Breastfeeding Disorders in the Presence of Paternal Inhibition and Maternal Neglect

Parents&#39; psychological stress during the perinatal and neonatal periods continues to increase in an environment of declining birthrates, aging populations, and shrinking family sizes. The increase in child abuse and neglect cases, most likely by inexperienced and insufficiently knowledgeable parents, necessitates education on childcare and intervention techniques in nursing and midwifery training. In particular, attachment formation early in life between mother and infant is crucial. To accurately teach sensitive and comprehensive information on intervention techniques for mother-child attachment formation, realistic videos, and educational materials are necessary. Although pseudoeducational materials are available, they might be limited in explaining complex realism, particularly to support breastfeeding that involves both parents and child and that encourages interaction between the two.
In a previous study in a common marmoset (Callithrix jacchus) model, we experimentally controlled infant feeding and nurturing through 24 h of constant sensing and collected 1 month of quantitative data on psychological indices that possibly translated to psychological development. Age-dependent dynamic visualization of these data by multivariate analyses inferred causal relationships between early parental feeding and psychobiological rhythm formation. In the same primate model, we identified a spontaneous case of breastfeeding failure in which the father inhibited his neonatal infant&#39;s feeding and the mother appeared to abandon nurturing, leading to clinically significant weight loss in the infant.
Thus, we explored intervention techniques to promote mother-infant interaction. The mother was trained to allow the infant to spontaneously explore her breast. Initially, the mother refused to display the feeding pose potentially due to pain associated with breast engorgement. Massage was used to soften the breast and feeding was reintroduced. We hypothesize that activation of instinctive attachment formation mechanisms by encouraging spontaneity in each parent and child is the key to successful feeding intervention.

Here, we present a nonhuman primate model of mother-infant intervention for breastfeeding disorders in the presence of paternal inhibition and maternal neglect. The mate model video complements education to support primate and human caregivers with respect to infants with breastfeeding problems such as pain.

Parents&#39; psychological stress during the perinatal and neonatal periods continues to increase in an environment of declining birthrates, aging ...

Behavior

A Surgical Procedure for the Administration of Drugs to the Inner Ear in a Non-Human Primate Common Marmoset (Callithrix jacchus)

Hearing research has long been facilitated by rodent models, although in some diseases, human symptoms cannot be recapitulated. The common marmoset (Callithrix jacchus) is a small, easy-to-handle New World monkey which has a similar anatomy of the temporal bone, including the middle ear ossicular chains and inner ear to humans, than in comparison with that of rodents. Here, we report a reproducible, safe, and rational surgical approach to the cochlear round window niche for the drug delivery to the inner ear of the common marmoset. We adopted posterior tympanotomy, a procedure used clinically in human surgery, to avoid manipulation of the tympanic membrane that may cause conductive hearing loss. This surgical procedure did not lead to any significant hearing loss. This approach was possible due to the large bulla structure of the common marmoset, although the lateral semicircular canal and vertical portion of the facial nerve should be carefully considered. This surgical method allows us to perform the safe and accurate administration of drugs without hearing loss, which is of great importance in obtaining pre-clinical proof of concept for translational research.

A Surgical Procedure for the Administration of Drugs to the Inner Ear in a Non-Human Primate Common Marmoset Callithrix jacchus

We report a surgical method to administrate drugs to the inner ear of a non-human primate, the common marmoset (Callithrix jacchus), via the round window membrane.

Hearing research has long been facilitated by rodent models, although in some diseases, human symptoms cannot be recapitulated. The common marmoset ...

Medicine

Chronic Implantation of Whole-cortical Electrocorticographic Array in the Common Marmoset

Electrocorticography (ECoG) allows the monitoring of electrical field potentials from the cerebral cortex with high spatiotemporal resolution. Recent development of thin, flexible ECoG electrodes has enabled conduction of stable recordings of large-scale cortical activity. We have developed a whole-cortical ECoG array for the common marmoset. The array continuously covers almost the entire lateral surface of cortical hemisphere, from the occipital pole to the temporal and frontal poles, and it captures whole-cortical neural activity in one shot. This protocol describes a chronic implantation procedure of the array in the epidural space of the marmoset brain. Marmosets have two advantages regarding ECoG recordings, one being the homologous organization of anatomical structures in humans and macaques, including frontal, parietal, and temporal complexes. The other advantage is that the marmoset brain is lissencephalic and contains a large number of complexes, which are more difficult to access in macaques with ECoG, that are exposed to the brain surface.These features allow direct access to most cortical areas beneath the surface of the brain. This system provides an opportunity to investigate global cortical information processing with high resolutions at a sub-millisecond order in time and millimeter order in space.

We have developed a whole-cortical electrocorticographic array for the common marmoset that continuously covers almost the entire lateral surface of cortex, from the occipital pole to the temporal and frontal poles. This protocol describes a chronic implantation procedure of the array in the epidural space of the marmoset brain.

Electrocorticography (ECoG) allows the monitoring of electrical field potentials from the cerebral cortex with high spatiotemporal resolution. Recent ...

Neuroscience

Stereotaxic Surgery for Implantation of Microelectrode Arrays in the Common Marmoset (Callithrix jacchus)

Marmosets (Callithrix jacchus) are small non-human primates that are gaining popularity in biomedical and preclinical research, including the neurosciences. Phylogenetically, these animals are much closer to humans than rodents. They also display complex behaviors, including a wide range of vocalizations and social interactions. Here, an effective stereotaxic neurosurgical procedure for implantation of recording electrode arrays in the common marmoset is described. This protocol also details the pre- and postoperative steps of animal care that are required to successfully perform such a surgery. Finally, this protocol shows an example of local field potential and spike activity recordings in a freely behaving marmoset 1 week after the surgical procedure. Overall, this method provides an opportunity to study the brain function in awake and freely behaving marmosets. The same protocol can be readily used by researchers working with other small primates. In addition, it can be easily modified to allow other studies requiring implants, such as stimulating electrodes, microinjections, implantation of optrodes or guide cannulas, or ablation of discrete tissue regions.

Stereotaxic Surgery for Implantation of Microelectrode Arrays in the Common Marmoset Callithrix jacchus

This work presents a protocol to perform a stereotaxic, neurosurgical implantation of&#160;microelectrode arrays in the common marmoset. This method specifically enables electrophysiological recordings in freely behaving animals but can be easily adapted to any other similar neurosurgical intervention in this species (e.g., cannula for drug administration or electrodes for brain stimulation).

Marmosets (Callithrix jacchus) are small non-human primates that are gaining popularity in biomedical and preclinical research, including the ...

Development of an X-Y Electrode Stage for Precise Neural Recording in Marmosets

Electrophysiology of Laminar Cortical Activity in the Common Marmoset

The marmoset monkey provides an ideal model for examining laminar cortical circuits due to its smooth cortical surface, which facilitates recordings with linear arrays. The marmoset has recently grown in popularity due to its similar neural functional organization to other primates and its technical advantages for recording and imaging. However, neurophysiology in this model poses some unique challenges due to the small size and lack of gyri as anatomical landmarks. Using custom-built micro-drives, researchers can manipulate linear array placement to sub-millimeter precision and reliably record at the same retinotopically targeted location across recording days. This protocol describes the step-by-step construction of the micro-drive positioning system and the neurophysiological recording technique with silicon linear electrode arrays. With precise control of electrode placement across recording sessions, researchers can easily traverse the cortex to identify areas of interest based on their retinotopic organization and the tuning properties of the recorded neurons. Further, using this laminar array electrode system, it is possible to apply a current source density analysis (CSD) to determine the recording depth of individual neurons. This protocol also demonstrates examples of laminar recordings, including spike waveforms isolated in Kilosort, which span multiple channels on the arrays.

Author Spotlight: Optimizing Micro-Drive Systems for Precise Electrode Positioning in Marmoset

Custom-built micro-drives enable the sub-millimeter targeting of cortical recording sites with linear silicon arrays.

The marmoset monkey provides an ideal model for examining laminar cortical circuits due to its smooth cortical surface, which facilitates recordings with ...

Noninvasive EEG for Comparative Studies of Vocal Perception in Marmosets

Electroencephalography Measurements in Awake Marmosets Listening to Conspecific Vocalizations

Vocal communication plays a crucial role in the social interactions of primates, particularly in survival and social organization. Humans have developed a unique and advanced vocal communication strategy in the form of language. To study the evolution of human language, it is necessary to investigate the neural mechanisms underlying vocal processing in humans, as well as to understand how brain mechanisms have evolved by comparing them with those in nonhuman primates. Herein, we developed a method to noninvasively measure the electroencephalography (EEG) of awake nonhuman primates. This recording method allows for long-term studies without harming the animals, and, importantly, allows us to directly compare nonhuman primate EEG data with human data, providing insights into the evolution of human language. In the current study, we used the scalp EEG recording method to investigate brain activity in response to species-specific vocalizations in marmosets. This study provides novel insights by using scalp EEG to capture widespread neural representations in marmosets during vocal perception, filling gaps in existing knowledge.

Author Spotlight: Investigating Vocal Information Representation in Small Primates and Its Alteration by Psychiatric Disorders Using Noninvasive EEG

To study the evolution of language, comparing brain mechanisms in humans with those in nonhuman primates is important. We developed a method to noninvasively measure the electroencephalography (EEG) of awake animals. It allows us to directly compare EEG data between humans and animals for the long term without harming them.

Vocal communication plays a crucial role in the social interactions of primates, particularly in survival and social organization. Humans have developed a ...

Assessing Ovarian Cycle in Marmosets by Blood Collection and Hormone Analysis

Blood Sampling and Hormone Measurement for Determining the Stage in the Ovarian Cycle in Marmosets

Common marmosets are small New World monkeys. Since many of their biological mechanisms are similar to those of humans, marmosets are potentially useful for medical and human biology research across a range of fields, such as neuroscience, regenerative medicine, and development. However, there is a lack of literature describing methods for many basic experiments and procedures. Here, detailed methods for determining the levels of sex hormones (progesterone, estradiol, and chorionic gonadotropin) in marmosets are described. The measurement of these hormones enables the prediction of the stage in the ovarian cycle, which is typically 26-30 days in marmosets; accurate determination is essential for the harvesting of oocytes/zygotes at the correct time point and for the preparation of host females for the generation of genetically modified marmosets. 
Additionally, the measurement of sex hormone levels is useful for endocrinology, ethology, early development, and reproductive biology studies. This protocol provides a detailed description of the methods for blood sampling from the femoral vein, separation of plasma for hormone measurement, measuring chorionic gonadotropin levels using urine and plasma, resetting the ovarian cycle using injections of a prostaglandin F2&#945; analog to shorten and synchronize the cycle, and promoting follicular growth and ovulation by injecting follicle-stimulating hormone and chorionic gonadotropin. Using these protocols, the stages in the ovarian cycle can be determined for the timely collection of oocytes/zygotes.

Author Spotlight: Advancing Primatology Through Germ Cell Research and Genetic Modification Techniques

This protocol describes blood and urine sampling for measuring progesterone/estradiol and chorionic gonadotropin levels to determine the ovarian cycle stage. The hormone levels are used to predict and determine the timing of ovulation and hormones are injected to regulate the ovarian cycle and oocyte growth.

Common marmosets are small New World monkeys. Since many of their biological mechanisms are similar to those of humans, marmosets are potentially useful ...

Rodent Handling and Restraint Techniques

Source: Kay Stewart, RVT, RLATG, CMAR; Valerie A. Schroeder, RVT, RLATG. University of Notre Dame, IN&#160;
It has been demonstrated that even minimal handling of mice and rats is stressful to the animals. Handling for cage changing and other noninvasive procedures causes an increase in heart rate, blood pressure, and other physiological parameters, such as serum corticosterone levels. Fluctuations can continue for up to several hours. The methods of restraint required for injections and blood withdrawals also cause physiological changes that can potentially affect scientific data. Training in the proper handling of mice and rats is required to minimize the effects to the animals.1 Mice and rats can be restrained manually with restraint devices, or with chemical agents. Manual methods and the use of restraint devices are covered in this manuscript. All restraint methods include the process of lifting the animals from their home cage.

Proper Handling and Restraining Techniques of Rodents

Source: Kay Stewart, RVT, RLATG, CMAR; Valerie A. Schroeder, RVT, RLATG. University of Notre Dame, IN&#160;
It has been demonstrated that even minimal ...

Hand-Rearing Method for Infant Marmosets

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