Crying is a universal act in infancy and an essential signal that activates care-giving behavior. While the details of crying have been documented in terms of correlations with other infant attributes, virtually nothing is known about the neural basis of crying or why crying can be such a compelling stimulus for the listener. Our primary objective is to understand the nature of the “cry circuit,” that is the neural pathways that underlie cry production and cry perception. We undertake behavioral experiments in nonhuman primates to define the critical features of infant crying that promote care-giving, to search for acoustic markers of developmental status through acoustic analysis of cry sounds, and to identify familial traits, environmental influences, and neurological risk factors. We also perform functional neuroanatomical studies aimed at defining the neural populations activated during crying and cry perception.
The cry circuit in the brain
Newman, Aronoff, Bernhards; in collaboration with Soltis
A necessary step in understanding how the brain regulates crying and cry responding is to identify the neural circuits that mediate crying and the response to it. Our laboratory has used two species of neotropical primate, the squirrel monkey and the common marmoset, as subjects for our work. Both species have typical primate brains, but their brain size readily lends itself to study detailed neuroanatomical structure under the microscope over a large part of the brain. We are also interested in the role of crying in activating and maintaining parenting behavior. To this end, we investigate the relationship between various hormones, such as prolactin, and parental behavior (Soltis et al., 2005). Future studies are aimed at understanding where and how prolactin and other hormones work at the level of specific neuronal populations to activate and maintain parental behavior.
Soltis J, Wegner FH, Newman JD. Urinary prolactin is correlated with mothering and allo-mothering in squirrel monkeys. Physiol Behav 2005;84:295-301.
Marmoset brain atlas
Newman, Aronoff; in collaboration with Bell, Silva
Any study of the neuroanatomy of the cry circuit requires knowledge of the structures are under investigation; a brain atlas serves this purpose. Despite the availability of two published brain atlases for the squirrel monkey, the only atlas published for the marmoset has been out of print for many years. Therefore, we undertook the task of creating a marmoset brain atlas. Using a digital camera, we photographed frontal (coronal) sections cut through the brain of an adult common marmoset and stained for cell bodies (Nissl stain) and fiber tracts (Weil stain); we used the resulting images for the atlas. Rachel Bell, working under our direction and that of Afonso Silva, used one of the existing squirrel monkey atlases as an aid in identifying and labeling the corresponding structures in the marmoset brain sections. To improve the usefulness of our atlas as a reference, we created schematic images of the photographed sections by using Adobe Photoshop and the NIH Image-J program. We labeled the schematic images rather than the actual photographs. In addition, we used a tablet monitor with an electronic stylus to create wire-frame tracings of the brain sections. With the use of MRI and functional MRI techniques, we will stretch or otherwise manipulate the tracings to overlay MRI images in planned studies of marmosets. The atlas, slated for publication, will contain both the photographs and the labeled schematic images.
Immunocytochemical identification of neurons active during crying
Newman, Aronoff, Bernhards, Rakhovskaya
Expression of the immediate-early gene c-fos has been used in a variety of animal models to label brain cells activated during the course of an extended period of a particular behavior. The procedure involves designing a behavioral protocol that promotes a particular definable form of behavior; allowing a period of time (typically 30 to 120 minutes) for gene activation, production of mRNA and the peptide product of the gene; and then sacrificing the animal for brain processing. The availability of an antibody specific for the c-fos peptide product Fos permits labeling of Fos-containing neurons by using immunocytochemistry techniques. Appropriate controls, including subjects not experiencing the selected behavioral protocol, ensure that Fos-labeled neurons are associated with the behavior under study. Crying is often a robust behavior that lasts for an extended period of time; hence, neurons activated during crying should be identifiable with the c-fos immunocytochemistry approach. Our studies in adult squirrel monkeys have confirmed that such is the case. Adult squirrel monkeys produce cry sounds very similar to those produced during infancy when individuals are separated from their social groups. Cry production is somewhat variable, with some individuals crying much more robustly than others. We have studied the brains of seven individuals, three robust vocalizers, three poor vocalizers, and one control animal (not separated from its social group). We used the Fos antibody to process free-floating sections cut coronally. Many labeled cells are visible under microscope at a 200x magnification in robust vocalizers, and far fewer labeled cells are visible in the poor vocalizers. To quantify the differences, we saved digitally photographed images and counted the labeled cells by using a commercially available program designed for this purpose. Among the more robust vocalizers, we found significantly higher numbers of labeled cells in the anterior cingulate gyrus, periventricular/periaqueductal gray matter, and anterior dorsal nuclei of the thalamus (which project to the anterior cingulate) and extensive labeling in the amygdala, claustrum, and hypothalamus, although we have not yet subjected the latter regions to quantified analysis.
Defining the cry circuit behaviorally
Newman, Bernhards, Aronoff; in collaboration with Soltis
Crying is the simplest form of a communication network, typically involving only the infant and its principal caregiver, often the mother. Furthermore, the acoustic structure of cry sounds is relatively stereotyped, making it possible to identify variations in acoustic structure that might reflect individual differences in brain development, early experience, and other early influences on crying. With growing interest in the common marmoset as a primate model for various neuropsychiatric disorders and its identification as a target species for complete genome analysis, researchers must determine which and to what extent marmoset behavioral patterns are under genetic control. The infant cry is a potentially important behavior in this effort, given its expression from birth and the evidence that individual experience plays little role in shaping the acoustic characteristics of cry sounds. Behavioral phenotyping, used successfully in the rhesus macaque and mice, is an important step in understanding gene-environment interactions, and crying provides the opportunity to study such interactions from the early neonatal period and onward.
Newman JD. Infant crying and colic: what lies beneath. Behav Brain Sci 2004;27:470-471.
Newman JD. The primate isolation call: a comparison with precocial birds and nonprimate mammals. In: Rogers L, Kaplan G, eds. Comparative Vertebrate Cognition: Are Primates Superior to Non-Primates? Dordrecht, Netherlands: Kluwer Academic Publishers, 2004:171-187.
Soltis J. The signal functions of early infant crying. Behav Brain Sci 2004;27:443-458.
Cry characteristics of infant marmosets
Newman, Bernhards, Aronoff, Ladd
For several years, we have relied on a standardized procedure for obtaining cry sounds from infant monkeys. The procedure involves gently separating an infant from its family group and taking it to a quiet room where no other monkeys or humans are present. Infants typically begin crying immediately, allowing us to obtain many cries in a short period. The infant is then returned to its family group. Our analysis of the acoustic structure of sounds made by crying infants identified one type of call in particular: the cry sound that persists throughout development (and into adulthood). To quantify developmental changes in crying and to assess the structural resemblance of cry sounds between genetically related and unrelated individuals, it is necessary to make detailed measurements of cry acoustic structure. To that end, we used a software program called “Raven,” developed at Cornell University, to measure the acoustic structure of the cry sounds of infant marmosets. We generated a sound spectrogram of each cry on a computer screen and saved the time and frequency values for four preselected points on each sound spectrogram. From the four time and frequency values, we calculated a total of 21 acoustic parameter measures by using a Microsoft Excel macro created by Karen Ladd; the tables of values are currently undergoing detailed statistical analysis. In addition, we have saved the parameter values for later statistical analysis.
COLLABORATORS
Rachel Bell (Senior, Montgomery Blair High School), Laboratory of Functional and Molecular Imaging, NINDS, Bethesda, MD
Afonso Silva, PhD, Laboratory of Functional and Molecular Imaging, NINDS, Bethesda, MD
Joseph Soltis, PhD, Disney’s Animal Kingdom, Lake Buena Vista, FL
For further information, contact newmanj@lce.nichd.nih.gov.