Studies have found that the complexity of songbirds' song was positively correlated with song nucleus, neuron density and neuron number (DeVoogd et al. 1993; Garamszegi and Eens 2004). The experimental resulted showed that there was no significant difference in the nuclear population volume and cell number of left and right HVC, It was proved that the endocrine system of songbirds played an equal and stable role in male birds with stable neurodevelopment in adulthood. Immunohistochemical results showed that the significant changes in songbirds' response to hormones may exist in the early stage of songbirds' song learning and development. Zhang et al. (2014) described that the right HVC had a significant advantage in the control of the frequency domain and intensity of singing through the click-damage experiment, but some studies have shown that the control of singing melody required the joint action of both HVC. It was not difficult to see that the right side of the song system was not an absolute control advantage over the song, but showed advantages in some signal output, that is to say, the output of song depended on the synergistic effect between the two sides of the song nuclei. Sound production depended on the auditory feedback, HVC was not only an important structure for song generation and learning, which was located at the highest level of singing motor pathway, but also a gated nucleus that received auditory information input (Nottebohm 1971), and integrate the new code was passed to give in to the front of the brain Are X or downstream muscle to produce the chirping of information feedback. The control level of HVC in the song system was higher than that of RA, but the nerve impulses generated by HVC could not directly control the activity of the song muscle. RA, as the efferent nucleus of HVC, was also the premotor nucleus that directly controlled the activity of nXII TS. RA could process neural signals from HVC, generated neural codes that could coordinate the interaction of the vocalis muscle and respiratory muscle, outputed to nXII TS and other related vocal motor nuclei to control singing behavior (Fee et al. 2004; Wang et al. 2018). There was no significant difference in nuclear population volume and cell number between left and right RA, but there was a difference in cell density. The cell density of right RA was significantly higher than that of left RA. If this result was caused by sample size and sampling error, the error would be further enlarged when the number of RA neurons was estimated, resulting in a significant difference in the number of neurons in left and right RA. However, the calculation results showed that there was no statistically significant difference in the number of RA cells (Fig. 6). So, another possible explanation of this phenomenon without considering the error was that the auditory processing and error feedback mechanism was biased to the right (Toyomura et al. 2007; Tourville et al. 2008), which leaded to the right HVC bearing more output of neural coding in the process of singing. Therefore, the right RA needs to integrate and output more regulatory information to regulate the movement of the vocalis muscle. On the other hand, In RA on the right, the number of nerve cells per unit volume increases in order to complete more complex information processing than that on the left, and more neurons and synaptic connections per unit volume can control the encoding of some special syllables in the process of singing. Right RA can accept more from the ipsilateral HVC and the lateral part of the striatum giant cell nucleus (LMAN), resulting in stronger regulation of the descending nucleus, which was consistent with the partial lateralization of its function. Whether right HVC has a stronger projection to ipsilateral RA has not been proved due to limited samples.
The production of complex songs were closely related to sexual selection, and males with more song editing ability were more likely to be approached by female songbirds (Spencer et al. 2005). This trait could be preserved and evolved under the restriction of genetic selection. Studies have shown that allometric relationships between ontogeny, static growth and developmental evolution were all influenced by a common factor, namely the growth process (Warton et al. 2006). Therefore, the degree of correlation between traits could be tested by estimating the covariance between relative traits. However, as the order of magnitude between traits may be different, phenotypic correlation coefficient was used in this experiment to compare multiple pairs of traits so that all parameters were compared at the same level. The growth rates of both sides HVC and RA were consistent with that of the brain (K > 1), indicating that in the upstream nucleus of the song learning and control system, the song control system had abundant variation among individuals, which was often beneficial to the evolution of the population, because the static growth relationship could often reflect the adaptation mode of individuals to environmental changes. Through experimental studies, Bondar et al. found that positive allometric growth seems to have a stronger inhibition on shape development in individuals (Meter et al. 2020). After comparing the correlation between HVC and RA, it was found that there was a higher correlation between right RA and HVC. What’s more, There was a positive allometric relationship between RA and HVC, It showed that the right RA in the process of development was the growth inhibition of mainly comes from the HVC’s three-dimensional shape, so as to brain size, but the ultimate factor in maintaining this relationship was natural selection. However, left RA showed negative allometric growth compared with HVC. It was generally believed that directional selection plays a role through the negative allometric growth between allometric growth. HVC and RA had a good fitness values and intercept was close with brain length, brain width and brain height. The singing nucleus always showed a negative growth relationship with three-dimensional parameters of the brain, which maybe because the brain always acts as a restriction to the neural nuclei occupying a certain spatial dimension, so as to improve the adaptability of individuals to environmental changes. Therefore, it was not difficult to find that the growth of the left and right nuclei was restricted by a variety of factors through static allometric growth of the singing nuclei. Among them, HVC was mainly influenced by natural selection, and there was mutual inhibition between HVC and ipsilateral RA, which was higher on the left than the right. The development process of RA had a certain delay on the individual level, so only the change of genetic covariance caused by natural selection or sexual preference for a long time can change its nuclear cluster development pattern. Therefore, when the living environment remained relatively constant, songbirds would continue to strictly follow the existing allometry growth pattern, and any individuals who seriously deviated from this development pattern would show more inadaptability. It was not hard to understand because, on an evolutionary scale, brain size's use of internal space required a measure of resource input and individual contribution, so the brain had always acted as a constraint on the number of neurons occupying a certain spatial dimension (Székely et al. 1996).