The thalamus is known to receive single-whisker 'lemniscal' inputs from the trigeminal nucleus principalis (PrV) and multiwhisker 'paralemniscal' inputs from the spinal trigeminal nucleus (SpV), yet the responses of cells in the thalamic ventroposteromedial nucleus (VPM) are most similar to and contingent upon inputs from PrV. This may reflect a differential termination pattern, density and/or synaptic organization of PrV and SpV projections. This hypothesis was tested in adult rats using anterograde double-labelling with fluorescent dextrans, horseradish peroxidase (HRP) and choleragenoid, referenced against parvalbumin and calbindin immunoreactivity. The results indicated that PrV's most robust thalamic projection is to the whisker-related barreloids of VPM. The SpV had robust projections to non-barreloid thalamic regions, including the VPM 'shell' encapsulating the barreloid area, a caudal and ventral region of VPM that lacks barreloids and PrV inputs, the posterior thalamic nucleus, nucleus submedius and zona incerta. Within the barreloid portion of VPM, SpV projections were sparse relative to those from PrV, and most terminal labelling occurred in the peripheral fringes of whisker-related patches and in interbarreloid septae. Thus, PrV and SpV have largely complementary projection foci in the thalamus. Intra-axonal staining of a small sample of trigeminothalamic axons with whisker or guard hair receptive fields revealed highly localized and somatotopic terminal aggregates in VPM that spanned areas no larger than that of a single barreloid. In the electron microscopic component of this study, HRP transport to the barreloid region of VPM from left SpV and right PrV in the same cases revealed PrV terminals contacting dendrites with a broad range of minor axis diameters (mean +/- SD: 1.51 +/- 0.10 microns). SpV terminals were indistinguishable from those of PrV, but they had a disproportionate number of contacts on narrow dendrites (1.27 +/- 0.07 microns, P < 0.01). PrV endings were also more likely to contact VPM somata (11.0 +/- 4.2% of all labelled terminals) than those from SpV (3.0 +/- 1.0%, P < 0.01). Insofar as primary dendrites are thicker than distal dendrites in VPM, these data suggest a differential distribution of PrV and SpV inputs onto VPM cells that may account for their relative efficacies in dictating the responses of VPM cells to whisker stimulation. Multiwhisker receptive fields in VPM may also reflect direct transmission of convergent inputs from PrV.

Nucleus HVC of the avian song system is essential to song patterning and is a prime site for auditory-vocal integration important to vocal learning. These processes require precise, high-frequency action potential activity, which, in other systems, is often correlated with the expression of calcium-binding proteins. To characterize any such functional specializations in HVC, we retrogradely labeled projection neurons innervating HVC's known targets, namely, area X or nucleus robustus arcopallialis (RA), then stained HVC sections with antibodies to the calcium-binding proteins parvalbumin, calbindin, and calretinin. Under epifluorescent illumination, neither projection neuron type exhibited detectable levels of calcium-binding protein immunoreactivity, whereas a third cell type, made up of nonprojection neurons (interneurons), was immunopositive for one, two, or all three of the calcium-binding proteins. In fact, most of these interneurons were either doubly or triply labeled. To explore the link between the electrical and calcium-binding protein properties of individual HVC neurons, we used intracellular methods in brain slices to record from identified HVC cell types based on their intrinsic electrical properties. Intracellular neurobiotin combined with immunostaining revealed that fast-spiking interneurons, but not the slower-spiking projection neurons, were positive for one or more calcium-binding proteins. Confocal microscopy confirmed these results and also revealed that RA-projecting cells might contain very low levels of parvalbumin. These results indicate that HVC interneurons are specialized in their calcium-binding proteins and suggest how it might be possible to resolve the details of HVC microcircuits underlying song selectivity and auditory-vocal learning.

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Ipsilateral and contralateral projections of nucleus robustus archistriatalis (RA), a telencephalic vocal premotor nucleus, to respiratory-vocal nuclei in the brainstem were defined in adult male Wasserschlager canaries, grey catbirds, and zebra finches, three songbird species that appear to differ in the degree of lateralized syringeal dominance. In all three species, ipsilateral projections of RA to the medulla included the tracheosyringeal part of the hypoglossal nucleus (XIIts), that innervates the syrinx, the bird's vocal organ, the suprahypoglossal area (SH), and two respiratory-related nuclei, retroambigualis (RAm) and parambigualis (PAm; Reinke and Wild [1998] J Comp Neurol 391:147-163). Projections of RA to the contralateral XIIts, SH and RAm, were substantial in canaries, which use the left side of the syrinx predominantly during singing; less pronounced in catbirds, which have no lateral dominance for song control; and least pronounced in zebra finches, in which there is a right-sided dominance for song control. There were no obvious differences in the number of crossed projections in birds injected in the left or right RA. Local sources of inputs to XIIts and RAm were defined anatomically in zebra finches and canaries. RAm, including neurons in close proximity to XIIts, was found to project to XIIts and the suprahypoglossal area bilaterally but predominantly ipsilaterally. RAm also had reciprocal connections with its contralateral homologue. These results suggest a pattern of connections between premotor and motor respiratory-vocal nuclei that may be involved in bilateral control of vocal output at medullary levels, a control that involves a high degree of coordination between vocal and respiratory structures on both sides of the body.