Harmonicity is a property of complex sounds such as vocalizations or music, but it remains unclear how harmonicity is processed in the auditory cortex (ACtx). Subregions of ACtx are thought to process harmonic stimuli differently. Selective responses to sound features in ACtx emerge hierarchically from primary ACtx (A1) L4 and secondary ACtx (A2) layer (L) 2/3, which is believed to be the most responsive to harmonic sounds. Since harmonic stacks can range from two to more than ten components, being more similar to naturalistic vocalizations, harmonic sensitivity might also arise hierarchically across layers and areas. We studied responses to harmonic stacks of two to ten frequencies across A1 L4, A1 L2/3, and A2 L2/3 in adult male and female mice using in vivo two-photon microscopy. We found harmonic-sensitive neurons (HN) responding only to harmonic stacks but not to individual frequencies in all areas at similar proportions. HNs showed highly nonlinear spectral integration of harmonic frequencies that decreased as the harmonic stacks became more complex. Specifically, onset-biased HNs showed greater nonlinearity than offset-biased HNs only in A1 L4. Moreover, HNs in A1 L4 exhibited higher signal correlation than A2 L2/3. Sound-responsive neurons in A1 L4 has weakest noise correlation compared to A1 L2/3 and A2 L2/3. Together, harmonic sensitivity is not a unique feature of A2 L2/3 but is already established in A1 L4, where neurons robustly encode harmonic sounds through sparse connections. Significance statement Harmonics are essential in auditory perception, influencing how we process complex sounds like music and speech. This study reveals that neurons in the cortical layer 4 and layer 2/3 integrate simple and complex harmonic structures with distinct mechanisms of neuronal recruitment. A1 L4 harmonic-sensitive neurons (HNs) demonstrated strong responses through high signal correlation and weak noise correlation, suggesting a robust but sparse mechanism for spectral integration. Our results show that harmonic relationships are already extracted at the input layers of A1, and that HNs show non-linear facilitative integration. Thus, tuning to sounds of complex spectral contents might be a fundamental processing function of the auditory cortex and is already established in A1 L4, which receives major thalamic inputs.
(陈韵如) et al. (Fri,) studied this question.