Rapid contact call-driven induction of NR2A and NR2B NMDA subunit mRNAs in the auditory thalamus of the budgerigar (Melopsittacus undulatus).

In situ hybridization histochemistry was used to assess the effect of auditory stimulation with natural contact calls on expression of NR2A and NR2B NMDA subunit mRNAs in neurons of the thalamic auditory relay nucleus ovoidalis (Ov) of a vocal learning parrot species, the budgerigar (Melopsittacus undulatus). The results showed that both the core (Ov) and ventromedial shell subdivisions (Ovm) of ovoidalis contained neurons expressing NR2A and NR2B mRNA in no-stimulation control subjects and that the distributions of neurons expressing these subunit mRNAs were very similar in both the core and shell of Ov. Contact call stimulation (5, 30 and 180 min) resulted in substantial increases of 50-60% in the number of neurons expressing NR2A and NR2B mRNAs in both the core and shell. Staining intensity, as measured by the optical density of stained somata approximately doubled compared to controls for both NR2 subunits in the 5 and 30 min conditions, but declined from 30 to 180 min. In all conditions, the density, but not staining intensity, of neurons expressing NR2B exceeded NR2A expression. Furthermore, the density of neurons expressing both subunit mRNAs in call stimulation conditions was greater in the core than in the shell despite the fact that total neuronal density was approximately 20% higher in the shell. Previous experiments have shown that call stimulation is more effective at inducing expression of the immediate early gene zenk in the Ov shell than core; however the present results do not indicate that either NR2A or NR2B mRNA expression mediates this effect since neither subunit exhibits greater expression in Ovm. Ca(++) release is needed for immediate early gene expression, however and, notably, Ovm contains large numbers of neurons containing CGRP, a peptide which has been shown to increase cytosolic Ca(++) levels.

Contact-call driven and tone-driven zenk expression in the nucleus ovoidalis of the budgerigar (Melopsittacus undulatus).

The effectiveness of species-typical contact calls and a 3-kHz pure tone to induce zenk gene protein expression in the primary thalamic auditory relay nucleus ovoidalis was compared in budgerigars (Melopsittacus undulatus), a parrot species capable of lifelong vocal learning. Ovoidalis consists of a core which projects topographically to field L of the telencephalon and a ventromedial shell containing many calcitonin-gene-related peptide neurons that project throughout field L as well as to an adjacent field receiving visual input. Tone-induced and call-induced zenk expression in the ovoidalis core were similar; however, call-induced zenk expression in ventromedial ovoidalis shell was significantly greater than tone-induced expression. These results support the idea that the ovoidalis shell may contain neurons specialized to process complex sounds including species-typical communication sounds.

Feeding and contact call stimulation both induce zenk and cfos expression in a higher order telencephalic area necessary for vocal learning in budgerigars.

Stimulation with natural contact calls and feeding were used to assess zenk and fos protein expression in budgerigars (Melopsittacus undulatus), a vocal learning parrot species in which feeding and physical contact often occur in conjunction with vocalization. Although only calls induced gene expression in Field L, the primary telencephalic auditory area, both calls and feeding induced gene expression in the frontal lateral nidopallium (NFl), a brain area in receipt of input from Field L which projects to areas afferent to vocal control nuclei and which is necessary for new call learning. NFl thus appears poised to provide both non-auditory as well as auditory feedback to the vocal system.

Sexual dimorphism of vocal control nuclei in budgerigars (Melopsittacus undulatus) revealed with Nissl and NADPH-d staining.

Nissl staining and nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) histochemistry were used to explore the existence of sexual dimorphism in vocal control nuclei of adult budgerigars (Melopsittacus undulatus), a parrot species capable of lifelong vocal learning. Behavioral studies indicate that adult males possess larger vocal repertoires than adult females and learn new calls more quickly. The results of the present study show that the volumes of all vocal nuclei, as measured using both Nissl-stained and NADPH-d-stained material, as well as the total numbers of NADPH-d neurons, were 35-110% greater in males. Furthermore, all vocal nuclei exhibit conspicuous NADPH-d staining compared to surrounding fields in both adult males and females. Nevertheless, there were no significant gender differences in either the intensity of neuropil staining or the densities of NADPH-d neurons in vocal nuclei. Moreover NADPH-d neuron somal shapes were similar in males and females. Diameters of NADPH-d neurons in vocal nuclei were 8.5-32% larger in males than in females. Greater size of NADPH-d neuronal somata in males may be a general property of this cell type in budgerigars because a similar gender difference was found in a visual nucleus, the entopallium, which is not directly associated with the vocal control system and does not exhibit sexual dimorphism in total volume or total NADPH-d neuron numbers. Taken together, the results of the present study favor the hypothesis that superior lifelong vocal learning ability in male budgerigars rests largely on larger volumes of vocal control nuclei in males rather than on sexual dimorphism in the internal composition of vocal nuclei.

Contact call-driven zenk mRNA expression in the brain of the budgerigar (Melopsittacus undulatus).

Contact call-driven zenk (zif268, egr1, NGF1A, Krox 24) mRNA expression was mapped with in situ hybridization histochemistry in a vocal learning parrot, the budgerigar (M. undulatus). Relative to controls, call stimulation induced high zenk mRNA expression in all auditory areas including those closely associated with the vocal system within the anterior forebrain (Brauth et al. (2001) J. Comp. Neurol. 432, 481; (2002) Learn. Memory 9, 76). Thus there is a high correspondence between the distributions of neurons exhibiting contact call-driven zenk protein and mRNA expression in budgerigars. Field L2a, an area reported previously to express only perinucleolar zenk protein localization (Brauth et al. (2002) Learn. Memory 9, 76) also showed zenk mRNA expression.

Contact call-driven Zenk protein induction and habituation in telencephalic auditory pathways in the Budgerigar (Melopsittacus undulatus): implications for understanding vocal learning processes.

Expression of the immediate early gene protein Zenk (zif 268, egr-1, NGF1A, Krox24) was induced in forebrain auditory nuclei in a vocal learning parrot species, the budgerigar (Melopsittacus undulatus), when the subjects either listened to playbacks of an unfamiliar contact call or to a contact call with which they had been familiarized previously. Auditory nuclei included the Field L complex (L1, L2a, and L3), the neostriatum intermedium pars ventrolateralis (NIVL), the neostriatum adjacent to caudal nucleus basalis (peri-basalis or pBas), an area in the frontal lateral neostriatum (NFl), the supracentral nucleus of the lateral neostriatum (NLs), and the ventromedial hyperstriatum ventrale (HVvm). The latter three nuclei are main sources of auditory input to the vocal system. Two patterns of nuclear staining were induced by contact call stimulation-staining throughout cell nuclei, which was exhibited by at least some neurons in all areas examined except L2a and perinucleolar staining, which was the only kind of staining exhibited in field L2a. The different patterns of Zenk staining indicate that auditory stimulation may regulate the Zenk-dependent transcription of different subsets of genes in different auditory nuclei. The numbers of neurons expressing Zenk staining increased from seven- to 43-fold over control levels when the birds listened to a repeating unfamiliar call. Familiarization of the subjects with the call stimulus, through repeated playbacks, greatly reduced the induction of Zenk expression to the call when it was presented again after an intervening 24-h interval. To determine if neurons exhibiting contact call-driven Zenk expression project to the vocal control system, call stimulation was coupled with dextran amines pathway tracing. The results indicated that tracer injections in the vocal nucleus HVo (oval nucleus of the hyperstriatum ventrale), in fields lateral to HVo and in NLs labeled many Zenk-positive neurons in HVvm, NFl, and NLs. These results support the idea that, in these neurons, egr-1 couples auditory stimulation to the synthesis of proteins involved in either the storing of new perceptual engrams for vocal learning or the processing of novel and/or meaningful acoustic stimuli related to vocal learning or the context in which it occurs.