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Partial inactivation of songbird auditory cortex impairs both tempo and pitch discrimination

Partial inactivation of songbird auditory cortex impairs both tempo and pitch discrimination Neuronal tuning for spectral and temporal features has been studied extensively in the auditory system. In the audi‑ tory cortex, diverse combinations of spectral and temporal tuning have been found, but how specific feature tuning contributes to the perception of complex sounds remains unclear. Neurons in the avian auditory cortex are spatially organized in terms of spectral or temporal tuning widths, providing an opportunity for investigating the link between auditory tuning and perception. Here, using naturalistic conspecific vocalizations, we asked whether subregions of the auditory cortex that are tuned for broadband sounds are more important for discriminating tempo than pitch, due to the lower frequency selectivity. We found that bilateral inactivation of the broadband region impairs performance on both tempo and pitch discrimination. Our results do not support the hypothesis that the lateral, more broadband subregion of the songbird auditory cortex contributes more to processing temporal than spectral information. Keywords Auditory perception, Auditory processing, Spectrotemporal receptive fields, Songbirds, Operant behavior Spectrotemporal receptive fields (STRFs) describe fun - perceptual tests with manipulation of neurons with a damental tuning properties of auditory neurons. Diverse particular STRF type may allow us to determine how dif- shapes of STRFs have been observed in cortical neu- ferently-tuned neurons contribute to the perception of a rons across different species [1, 2]. It has been hypoth- complex sound. esized that the different types of STRFs underlie basic In the songbird auditory cortex field L, neurons are sound percepts such as pitch and tempo [3]. For exam- tuned for spectrotemporal modulations of learned songs ple, broadband neurons may primarily extract temporal [4] and receptive fields have been categorized into func - changes in sound intensity, whereas narrow band neu- tional groups based on spectral and temporal tuning rons may be better at extracting spectral frequency infor- properties [3, 5, 6]. Moreover, STRFs are anatomically mation. However, whether a specific type of tuning con - organized in field L, with spectral tuning widths broad - tributes to the perception of complex sounds, including ening laterally along the mediolateral axis, and temporal learned vocalizations, remains to be tested. Combining tuning widths broadening along the dorsoventral axis from input to output layers [7]. In the thalamic input- receiving layer, a gradient of spectral tuning widths *Correspondence: exist, in which spectral tuning broadens laterally, while Gunsoo Kim kgunsoo@kbri.re.kr temporal tuning remains narrow. The spectral broaden - Sensory and Motor Systems Research Group, Korea Brain Research ing is also observed in the output layers despite a sharp Institute, Daegu, South Korea 2 increase in temporal tuning widths. Therefore, the lat - Department of Biology, Tufts University, Medford, MA 02155, USA Graduate School of Biomedical Sciences, Tufts University School eral subregion, being less selective for specific frequen - of Medicine, Boston, MA 02111, USA cies across layers, may be more important for processing © The Author(s) 2023. Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http:// creat iveco mmons. org/ licen ses/ by/4. 0/. The Creative Commons Public Domain Dedication waiver (http:// creat iveco mmons. org/ publi cdoma in/ zero/1. 0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data. Kim et al. Molecular Brain (2023) 16:48 Page 2 of 5 temporal information than spectral information. Prior approximately 1–3 mm lateral to the midline, sparing a studies have shown that songbirds pay close attention substantial medial portion of field L  of the infused solu - to conspecific vocalizations, making them an excellent tions (Fig. 1G; see Additional file 1: Methods). model for behavioral investigations of auditory percep- Birds’ performance across different conditions was tion [8–10]. As a first step towards linking specific tun - quantified as the probability of responding “fast” or ing to perception, we pharmacologically inactivated the “high” for each tempo or pitch shift (Fig. 1E and F). In the lateral part of the auditory cortex while songbirds per- tempo group, during bilateral infusion of phosphate buff - formed auditory discrimination tasks. ered saline (PBS; 0.025 M), performance did not differ Using a two-alternative forced choice task, adult from pre-surgery sessions (Fig.  1C and E; p values > 0.8 female zebra finches, who do not sing but do discrimi - (pre-surgery vs. PBS) for all tempo shifts, Tukey–Kramer nate between and show preferences for particular songs post hoc tests, following ANOVA with pre-surgery, PBS, and calls [8–11], were trained to discriminate modified and muscimol conditions). During bilateral muscimol zebra finch songs based on tempo or pitch (4 birds in infusion, performance on tempo discrimination was sig- the tempo group and 4 in the pitch group). In the tempo nificantly reduced compared to the PBS condition (from task, five renditions of a conspecific song were sped up or 91 ± 1% to 69 ± 2%, n = 4; Fig.  1C and E; p < 0.0005 for slowed down (± 16%, ± 8%, ± 4%), while maintaining the all shifts (PBS vs. bilateral muscimol), Tukey–Kramer original pitch. In the pitch task, five renditions of a differ - post-hoc tests corrected for multiple comparison; see ent conspecific song were shifted up or down in overall Additional file  2: Fig. S2A for individual performance), pitch (± 1/12, ± 1/24, ± 1/60 octaves), while maintaining although it did remain above chance. Performance recov- the original tempo (Fig. 1A, B; [8]). All stimuli were mod- ered upon switching back to PBS. These results indicate ified songs and presented in a pseudorandom manner. that the transient inactivation of the broadband, lateral The tempo group was trained to categorize a song as fast subregion of field L can impair birds’ ability to discrimi - or slow by pressing one of the two response perches, and nate song stimuli based on their tempo. the pitch group was trained to categorize a song as high If spectrally broad lateral subregions are specialized or low. Birds were rewarded with food in correct trials for tempo, inactivating these regions should not disrupt and punished in incorrect trials by a lights out period pitch discrimination. During bilateral muscimol infusion during which all perches stopped working temporarily. in the lateral subregion, however, the pitch group’s overall Birds learned the tasks over a period of weeks performance also dropped significantly [from 87 ± 2% to (Fig.  1C, D), reaching a performance plateau (> 70% 74 ± 1%; n = 4; Fig. 1D and F; p < 0.05 for − 1/12, − 1/24, − correct responses) first with the stimuli with larger 1/60, and + 1/12 (PBS vs. bilateral muscimol)], although shifts and later with stimuli with smaller shifts (tempo it still remained above chance. The reduction in perfor - task (mean ± SEM days): 17.5 ± 0.5 days on 16% shifts mance was smaller in the pitch group compared to the vs. 34.8 ± 5.5 days on 4% shifts, n = 4 birds; pitch task: tempo group, and for some pitch shifts, performance 12.0 ± 2.7 days on 1/12octave shifts vs. 22.5 ± 3.6 days on during muscimol was not significantly different from that 1/60 octave shifts, n = 4 birds). Once they became profi - during PBS infusion (+ 1/60 oct (p = 0.25) and + 1/24 oct cient at the tasks, performance was maintained at > 70% (p = 0.17); see Additional file  2: Fig. S2B for individual for all tempo and pitch shifts (Additional file  2: Fig. S1). performance). Nonetheless, the muscimol-induced per- Mean response latencies measured from the stimulus off - formance drop in the pitch group does not support the set were 0.65 ± 0.06 s for the tempo group and 0.76 ± 0.1 s hypothesis that the lateral subregion of field L is special - for the pitch group (p = 0.56, two sample t test). ized for tempo processing. After each bird learned to discriminate songs based In both groups, response rates and latencies did not on pitch or tempo, cannulae were implanted and micro- differ significantly across pre-surgery, PBS, and mus - dialysis probes were inserted in the lateral subregion of cimol conditions, indicating the implantation surgery field L of both hemispheres to manipulate the activity of itself did not cause gross impairments (“tempo group”: neurons with broadband spectral tuning widths [12]. We response rate (mean ± SE M): 81 ± 3% (pre-surger y), examined the effects of inactivating the cortical subregion 88 ± 4% (PBS), 83 ± 4% (muscimol), F(2,9) = 1.15, with broader spectral tuning [7] on tempo discrimination p = 0.36, ANOVA; latency (mean ± SE M): 0.65 ± 0.06 and pitch discrimination by reverse-dialyzing muscimol, s (pre-surgery), 0.59 ± 0.05 s (PBS), 0.64 ± 0.03 s a potent GABA receptor agonist (1 mM). Muscimol can (muscimol), F(2,9) = 0.29, p = 0.92, ANOVA; “pitch inactivate neurons near the infusion site without affect - group”: response rate: 78 ± 7% (pre-surgery), 76 ± 9% ing the fibers of passage and has been used successfully in (PBS), 81 ± 3% (muscimol), F(2,9) = 0.13, p = 0.88, songbirds [12, 13]. Our histology suggests that the musci- ANOVA; latency: 0.76 ± 0.1 s (pre-surgery), 0.69 ± 0.1 mol infusion was concentrated in lateral field L, spanning s (PBS), 0.62 ± 0.07 s (muscimol), F(2,9) = 0.1, p = 0.91, K im et al. Molecular Brain (2023) 16:48 Page 3 of 5 Δtempo Δpitch AB High Fast (+1/12 oct) (+16%) (-16%) (-1/12 oct) -1/12 oct +1/12 oct Slow Low 0.5 sec Tempo example bird Pitch example bird CD Fast High saline muscimol (bilateral) Implant Implant muscimol (unilateral) Slow Low Implant Implant Blocks Blocks Medial Lateral EF Tempo group (n = 4 birds) Pitch group (n = 4 birds) G 0.51.0 1.52 ** ****** CM CM L CM CM *** ****** **** ** Slow Fast Low High Fig. 1 Bilateral muscimol infusion in lateral field L lowers performance on both tempo and pitch discrimination tasks. A, B Example song stimuli for tempo (A) and pitch (B) discrimination tasks. Only the stimuli with largest tempo (± 16%) and pitch shifts (± 1/12 octaves) are shown. Inset in B Spectrogram of an example syllable (green boxes) to illustrate pitch shifts (numbers 1 to 5 label corresponding harmonics between the two shifts). C, D Example learning curves for one bird trained on the tempo task (C) and a different bird trained on the pitch task (D). Data are grouped into blocks of 200 trials (~ 3 blocks / day). The lines with different shades of gray (black, dark gray, and light gray) represent 3 different shifts in tempo or pitch with lighter lines representing smaller shifts. The triangles on the x‑axis indicate the time of implant surgery. The horizontal bars indicate infusion sessions that occurred on different days (blue: bilateral PBS; red: bilateral muscimol; green: unilateral muscimol). E, F Psychometric curves for different infusion sessions (blue: PBS; red: bilateral muscimol; green: unilateral muscimol; mean ± SEM). *p < 0.05, **p < 0.01, ***p < 0.001, Tukey–Kramer post‑hoc test (PBS vs. bilateral muscimol). No significant difference found between PBS and unilateral muscimol condition. G Photomicrographs of auditory forebrain regions in parasagittal sections that show the extent of biotinylated muscimol spread from ~ 0.5 mm lateral, where no biotin staining is visible, to ~ 2 mm lateral, where a dialysis probe was implanted. Top rows: 40 μm sections stained for biotinylated muscimol and adjacent Nissl stained sections. Bottom plot: the intensity of biotin staining as a function of distance from the midline (1 hemisphere from a tempo bird; 8 hemispheres from 4 pitch birds). The thick black line is from the example sections shown above. L: field L; CM: caudal mesopallium. Scale bar = 1 mm NisslBiotin Kim et al. Molecular Brain (2023) 16:48 Page 4 of 5 ANOVA). We also did not observe significant changes even cell types [18], could reveal relationships between in performance during unilateral infusion of muscimol spatially segregated sound feature encoding in the audi- in both groups (Fig.  1C–F; tempo: p > 0.2, n = 3 birds; tory cortex and the processing of fundamental percepts pitch: p > 0.7, n = 4 birds, Tukey–Kramer post-hoc tests, of complex sounds. following ANOVA with pre-surgery, PBS, unilateral and bilateral muscimol conditions). Abbreviations In this study, we investigated the link between auditory STRF Spectrotemporal receptive fields tuning properties and perception of complex sounds. We PBS Phosphate buffered saline asked whether the lateral subregion of the songbird pri- mary auditory cortex, which exhibits broadband spec- Supplementary Information tral tuning, contributes specifically to tempo processing, The online version contains supplementary material available at https:// doi. org/ 10. 1186/ s13041‑ 023‑ 01039‑5. without affecting pitch discrimination. Our results show that reversible bilateral inactivation of lateral field L Additional file 1. Methods. caused a significant reduction in performance in both Additional file 2: Figure S1. Performance comparison across different tempo and pitch discrimination. Therefore, while prior magnitudes of tempo and pitch shift before implant surgery. (A–B) Prob‑ work has shown that the spectrotemporal receptive fields ability of correct trials for different tempo (A) or pitch shifts (B) (mean ± SEM). Different shades of gray for bars indicate difficulty and correspond of neurons in the songbird auditory cortex are spatially to those in Fig. 1C and D. Performance was not significantly different organized, our results do not provide strong support for across different magnitude of shifts for either group (tempo group: F(5,18) the hypothesis that different subregions of auditory cor - = 2, p = 0.13; pitch group: F (5,18) = 2.64, p = 0.059, ANOVA). Figure S2. Performance of individual birds during bilateral muscimol infusion. (A–B) tex subserve different percepts. Probability of correct trials for different tempo (A) and pitch shifts (B). The Several limitations affect our ability to make strong blue and red lines show the average (± SEM) P(correct) for saline and conclusions about whether different subregions of field bilateral muscimol conditions, respectively (n = 4 birds on tempo task; n = 4 birds on pitch task). The gray lines show P (correct) for individual birds L differentially contribute to basic perceptual qualities during bilateral muscimol infusions, and the dashed lines indicate the per‑ of sound. First, we did not perform the converse experi- formance of the two example birds shown in Fig. 1C and D, respectively. ment—transient inactivation of the medial subregion of Field L—to test whether it differentially affects pitch vs. Acknowledgements tempo processing. Second, our muscimol infusion in We thank Adria Arteseros and Hyesook Lee for technical assistance. the lateral field L spread dorsoventrally across all lay - Author contributions ers of Field L and part of the caudal mesopallium (CM), GK conceived and performed experiments, and analyzed data. MS‑ V imaged a secondary cortical region, affecting neurons that may and analyzed histological data. GK, MS‑ V, and MHK wrote the manuscript. All play different roles in tempo and pitch processing due authors read and approved the final manuscript. to their markedly different temporal tuning properties Funding [7, 14, 15]. Third, we did not perform neural recordings This work was supported by NIH Grant R03DC012428, KBRI basic research during muscimol infusion to confirm whether musci - program through Korea Brain Research Institute funded by Ministry of Science and ICT of Korea (23‑BR‑01‑01, 23‑BR‑04‑04, and 22‑BR‑03‑07), and Basic mol inactivated lateral neurons while sparing the tun- Research Program of the National Research Foundation of Korea funded by ing properties of medial neurons. Although the above the Ministry of Science and ICT of Korea (NRF‑2021R1F1A1049434) to G.K, chance level performance during muscimol infusion Korea Brain Research Institute funded by Ministry of Science and ICT of Korea (23‑BR‑02‑10) to M.S‑ V, NIH grant R01DC011356 to Allison Doupe, and NIH and our histological quantification (Fig.  1G) indicate Grant R01NS129695 to M.H.K. that a substantial medial portion of field L was spared of muscimol, neural recordings would provide more Availability of data and materials All data and materials are available from the corresponding author upon direct examination of the muscimol effect, includ - reasonable request. ing potential changes in tuning via local connections. Finally, while a global gradient in the receptive fields Declarations of neurons in field L has been observed in songbirds, more recent studies in birds and mammals have found Ethics approval and consent to participate All procedures were performed in accordance with protocols approved by local heterogeneity in response properties of auditory the University of California, San Francisco Institutional Animal Care and Use neurons [16, 17], raising the possibility that local het- Committee (AN107972). erogeneity in spectral tuning widths may counteract the Consent for publication effect of inactivation based on a global gradient. Future Not applicable. experiments with a better spatial control of neural activity, such as optogenetic manipulations combined Competing interests The authors declare that they have no competing interests. with genetic targeting of specific regions, layers, or K im et al. Molecular Brain (2023) 16:48 Page 5 of 5 Received: 29 December 2022 Accepted: 25 May 2023 References 1. Eggermont JJ, Johannesma PM, Aertsen AM. Reverse‑ correlation meth‑ ods in auditory research. Q Rev Biophys. 1983;16:341–414. 2. Theunissen FE, Sen K, Doupe AJ. Spectral‑temporal receptive fields of nonlinear auditory neurons obtained using natural sounds. J Neurosci. 2000;20:2315–31. 3. Woolley SMN, Gill PR, Fremouw T, Theunissen FE. Functional groups in the avian auditory system. J Neurosci. 2009;29:2780–93. 4. Moore JM, Woolley SMN. Emergent tuning for learned vocalizations in auditory cortex. Nat Neurosci. 2019;22:1469–76. 5. Nagel KI, Doupe AJ. Organizing principles of spectro‑temporal encoding in the avian primary auditory area field L. Neuron. 2008;58:938–55. 6. Nagel K, Kim G, McLendon H, Doupe A. A bird brain’s view of auditory processing and perception. Hear Res. 2011;273:123–33. 7. Kim G, Doupe A. Organized representation of spectrotemporal features in songbird auditory forebrain. J Neurosci. 2011;31:16977–90. 8. Nagel KI, McLendon HM, Doupe AJ. Differential influence of frequency, timing, and intensity cues in a complex acoustic categorization task. J Neurophysiol. 2010;104:1426–37. 9. Elie JE, Theunissen FE. Zebra finches identify individuals using vocal signatures unique to each call type. Nat Commun. 2018;9:1–11. 10. Woolley SC, Doupe AJ. Social context‑induced song variation affects female behavior and gene expression. PLoS Biol. 2008;6: e62. 11. Paul A, McLendon H, Rally V, Sakata JT, Woolley SC. Behavioral discrimi‑ nation and time‑series phenotyping of birdsong performance. PLoS Comput Biol. 2021;17: e1008820. 12. Moorman S, Ahn J‑R, Kao MH. Plasticity of stereotyped birdsong driven by chronic manipulation of cortical‑basal ganglia activity. Curr Biol. 2021;31:2619‑2632.e4. 13. Tanaka M, Singh Alvarado J, Murugan M, Mooney R. Focal expression of mutant huntingtin in the songbird basal ganglia disrupts cortico‑ basal ganglia networks and vocal sequences. Proc Natl Acad Sci. 2016;113:E1720–7. 14. Lim Y, Lagoy R, Shinn‑ Cunningham BG, Gardner TJ. Transforma‑ tion of temporal sequences in the zebra finch auditory system. Elife. 2016;5:e18205. 15. Calabrese A, Woolley SMN. Coding principles of the canonical cortical microcircuit in the avian brain. Proc Natl Acad Sci. 2015;112:3517–22. 16. Araki M, Bandi MM, Yazaki‑Sugiyama Y. Mind the gap: neural coding of species identity in birdsong prosody. Science. 2016;354:1282–7. 17. Zeng H, Huang J, Chen M, Wen Y, Shen Z, Poo M. Local homogeneity of tonotopic organization in the primary auditory cortex of marmosets. Proc Natl Acad Sci. 2019;116:3239–44. 18. Lee C, Lavoie A, Liu J, Chen SX, Liu B. Light up the brain: the application of optogenetics in cell‑type specific dissection of mouse brain circuits. Front Neural Circuits. 2020. https:// doi. org/ 10. 3389/ fncir. 2020. 00018. Publisher’s Note Springer Nature remains neutral with regard to jurisdictional claims in pub‑ lished maps and institutional affiliations. Re Read ady y to to submit y submit your our re researc search h ? Choose BMC and benefit fr ? Choose BMC and benefit from om: : fast, convenient online submission thorough peer review by experienced researchers in your field rapid publication on acceptance support for research data, including large and complex data types • gold Open Access which fosters wider collaboration and increased citations maximum visibility for your research: over 100M website views per year At BMC, research is always in progress. Learn more biomedcentral.com/submissions http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Molecular Brain Springer Journals

Partial inactivation of songbird auditory cortex impairs both tempo and pitch discrimination

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Abstract

Neuronal tuning for spectral and temporal features has been studied extensively in the auditory system. In the audi‑ tory cortex, diverse combinations of spectral and temporal tuning have been found, but how specific feature tuning contributes to the perception of complex sounds remains unclear. Neurons in the avian auditory cortex are spatially organized in terms of spectral or temporal tuning widths, providing an opportunity for investigating the link between auditory tuning and perception. Here, using naturalistic conspecific vocalizations, we asked whether subregions of the auditory cortex that are tuned for broadband sounds are more important for discriminating tempo than pitch, due to the lower frequency selectivity. We found that bilateral inactivation of the broadband region impairs performance on both tempo and pitch discrimination. Our results do not support the hypothesis that the lateral, more broadband subregion of the songbird auditory cortex contributes more to processing temporal than spectral information. Keywords Auditory perception, Auditory processing, Spectrotemporal receptive fields, Songbirds, Operant behavior Spectrotemporal receptive fields (STRFs) describe fun - perceptual tests with manipulation of neurons with a damental tuning properties of auditory neurons. Diverse particular STRF type may allow us to determine how dif- shapes of STRFs have been observed in cortical neu- ferently-tuned neurons contribute to the perception of a rons across different species [1, 2]. It has been hypoth- complex sound. esized that the different types of STRFs underlie basic In the songbird auditory cortex field L, neurons are sound percepts such as pitch and tempo [3]. For exam- tuned for spectrotemporal modulations of learned songs ple, broadband neurons may primarily extract temporal [4] and receptive fields have been categorized into func - changes in sound intensity, whereas narrow band neu- tional groups based on spectral and temporal tuning rons may be better at extracting spectral frequency infor- properties [3, 5, 6]. Moreover, STRFs are anatomically mation. However, whether a specific type of tuning con - organized in field L, with spectral tuning widths broad - tributes to the perception of complex sounds, including ening laterally along the mediolateral axis, and temporal learned vocalizations, remains to be tested. Combining tuning widths broadening along the dorsoventral axis from input to output layers [7]. In the thalamic input- receiving layer, a gradient of spectral tuning widths *Correspondence: exist, in which spectral tuning broadens laterally, while Gunsoo Kim kgunsoo@kbri.re.kr temporal tuning remains narrow. The spectral broaden - Sensory and Motor Systems Research Group, Korea Brain Research ing is also observed in the output layers despite a sharp Institute, Daegu, South Korea 2 increase in temporal tuning widths. Therefore, the lat - Department of Biology, Tufts University, Medford, MA 02155, USA Graduate School of Biomedical Sciences, Tufts University School eral subregion, being less selective for specific frequen - of Medicine, Boston, MA 02111, USA cies across layers, may be more important for processing © The Author(s) 2023. Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http:// creat iveco mmons. org/ licen ses/ by/4. 0/. The Creative Commons Public Domain Dedication waiver (http:// creat iveco mmons. org/ publi cdoma in/ zero/1. 0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data. Kim et al. Molecular Brain (2023) 16:48 Page 2 of 5 temporal information than spectral information. Prior approximately 1–3 mm lateral to the midline, sparing a studies have shown that songbirds pay close attention substantial medial portion of field L  of the infused solu - to conspecific vocalizations, making them an excellent tions (Fig. 1G; see Additional file 1: Methods). model for behavioral investigations of auditory percep- Birds’ performance across different conditions was tion [8–10]. As a first step towards linking specific tun - quantified as the probability of responding “fast” or ing to perception, we pharmacologically inactivated the “high” for each tempo or pitch shift (Fig. 1E and F). In the lateral part of the auditory cortex while songbirds per- tempo group, during bilateral infusion of phosphate buff - formed auditory discrimination tasks. ered saline (PBS; 0.025 M), performance did not differ Using a two-alternative forced choice task, adult from pre-surgery sessions (Fig.  1C and E; p values > 0.8 female zebra finches, who do not sing but do discrimi - (pre-surgery vs. PBS) for all tempo shifts, Tukey–Kramer nate between and show preferences for particular songs post hoc tests, following ANOVA with pre-surgery, PBS, and calls [8–11], were trained to discriminate modified and muscimol conditions). During bilateral muscimol zebra finch songs based on tempo or pitch (4 birds in infusion, performance on tempo discrimination was sig- the tempo group and 4 in the pitch group). In the tempo nificantly reduced compared to the PBS condition (from task, five renditions of a conspecific song were sped up or 91 ± 1% to 69 ± 2%, n = 4; Fig.  1C and E; p < 0.0005 for slowed down (± 16%, ± 8%, ± 4%), while maintaining the all shifts (PBS vs. bilateral muscimol), Tukey–Kramer original pitch. In the pitch task, five renditions of a differ - post-hoc tests corrected for multiple comparison; see ent conspecific song were shifted up or down in overall Additional file  2: Fig. S2A for individual performance), pitch (± 1/12, ± 1/24, ± 1/60 octaves), while maintaining although it did remain above chance. Performance recov- the original tempo (Fig. 1A, B; [8]). All stimuli were mod- ered upon switching back to PBS. These results indicate ified songs and presented in a pseudorandom manner. that the transient inactivation of the broadband, lateral The tempo group was trained to categorize a song as fast subregion of field L can impair birds’ ability to discrimi - or slow by pressing one of the two response perches, and nate song stimuli based on their tempo. the pitch group was trained to categorize a song as high If spectrally broad lateral subregions are specialized or low. Birds were rewarded with food in correct trials for tempo, inactivating these regions should not disrupt and punished in incorrect trials by a lights out period pitch discrimination. During bilateral muscimol infusion during which all perches stopped working temporarily. in the lateral subregion, however, the pitch group’s overall Birds learned the tasks over a period of weeks performance also dropped significantly [from 87 ± 2% to (Fig.  1C, D), reaching a performance plateau (> 70% 74 ± 1%; n = 4; Fig. 1D and F; p < 0.05 for − 1/12, − 1/24, − correct responses) first with the stimuli with larger 1/60, and + 1/12 (PBS vs. bilateral muscimol)], although shifts and later with stimuli with smaller shifts (tempo it still remained above chance. The reduction in perfor - task (mean ± SEM days): 17.5 ± 0.5 days on 16% shifts mance was smaller in the pitch group compared to the vs. 34.8 ± 5.5 days on 4% shifts, n = 4 birds; pitch task: tempo group, and for some pitch shifts, performance 12.0 ± 2.7 days on 1/12octave shifts vs. 22.5 ± 3.6 days on during muscimol was not significantly different from that 1/60 octave shifts, n = 4 birds). Once they became profi - during PBS infusion (+ 1/60 oct (p = 0.25) and + 1/24 oct cient at the tasks, performance was maintained at > 70% (p = 0.17); see Additional file  2: Fig. S2B for individual for all tempo and pitch shifts (Additional file  2: Fig. S1). performance). Nonetheless, the muscimol-induced per- Mean response latencies measured from the stimulus off - formance drop in the pitch group does not support the set were 0.65 ± 0.06 s for the tempo group and 0.76 ± 0.1 s hypothesis that the lateral subregion of field L is special - for the pitch group (p = 0.56, two sample t test). ized for tempo processing. After each bird learned to discriminate songs based In both groups, response rates and latencies did not on pitch or tempo, cannulae were implanted and micro- differ significantly across pre-surgery, PBS, and mus - dialysis probes were inserted in the lateral subregion of cimol conditions, indicating the implantation surgery field L of both hemispheres to manipulate the activity of itself did not cause gross impairments (“tempo group”: neurons with broadband spectral tuning widths [12]. We response rate (mean ± SE M): 81 ± 3% (pre-surger y), examined the effects of inactivating the cortical subregion 88 ± 4% (PBS), 83 ± 4% (muscimol), F(2,9) = 1.15, with broader spectral tuning [7] on tempo discrimination p = 0.36, ANOVA; latency (mean ± SE M): 0.65 ± 0.06 and pitch discrimination by reverse-dialyzing muscimol, s (pre-surgery), 0.59 ± 0.05 s (PBS), 0.64 ± 0.03 s a potent GABA receptor agonist (1 mM). Muscimol can (muscimol), F(2,9) = 0.29, p = 0.92, ANOVA; “pitch inactivate neurons near the infusion site without affect - group”: response rate: 78 ± 7% (pre-surgery), 76 ± 9% ing the fibers of passage and has been used successfully in (PBS), 81 ± 3% (muscimol), F(2,9) = 0.13, p = 0.88, songbirds [12, 13]. Our histology suggests that the musci- ANOVA; latency: 0.76 ± 0.1 s (pre-surgery), 0.69 ± 0.1 mol infusion was concentrated in lateral field L, spanning s (PBS), 0.62 ± 0.07 s (muscimol), F(2,9) = 0.1, p = 0.91, K im et al. Molecular Brain (2023) 16:48 Page 3 of 5 Δtempo Δpitch AB High Fast (+1/12 oct) (+16%) (-16%) (-1/12 oct) -1/12 oct +1/12 oct Slow Low 0.5 sec Tempo example bird Pitch example bird CD Fast High saline muscimol (bilateral) Implant Implant muscimol (unilateral) Slow Low Implant Implant Blocks Blocks Medial Lateral EF Tempo group (n = 4 birds) Pitch group (n = 4 birds) G 0.51.0 1.52 ** ****** CM CM L CM CM *** ****** **** ** Slow Fast Low High Fig. 1 Bilateral muscimol infusion in lateral field L lowers performance on both tempo and pitch discrimination tasks. A, B Example song stimuli for tempo (A) and pitch (B) discrimination tasks. Only the stimuli with largest tempo (± 16%) and pitch shifts (± 1/12 octaves) are shown. Inset in B Spectrogram of an example syllable (green boxes) to illustrate pitch shifts (numbers 1 to 5 label corresponding harmonics between the two shifts). C, D Example learning curves for one bird trained on the tempo task (C) and a different bird trained on the pitch task (D). Data are grouped into blocks of 200 trials (~ 3 blocks / day). The lines with different shades of gray (black, dark gray, and light gray) represent 3 different shifts in tempo or pitch with lighter lines representing smaller shifts. The triangles on the x‑axis indicate the time of implant surgery. The horizontal bars indicate infusion sessions that occurred on different days (blue: bilateral PBS; red: bilateral muscimol; green: unilateral muscimol). E, F Psychometric curves for different infusion sessions (blue: PBS; red: bilateral muscimol; green: unilateral muscimol; mean ± SEM). *p < 0.05, **p < 0.01, ***p < 0.001, Tukey–Kramer post‑hoc test (PBS vs. bilateral muscimol). No significant difference found between PBS and unilateral muscimol condition. G Photomicrographs of auditory forebrain regions in parasagittal sections that show the extent of biotinylated muscimol spread from ~ 0.5 mm lateral, where no biotin staining is visible, to ~ 2 mm lateral, where a dialysis probe was implanted. Top rows: 40 μm sections stained for biotinylated muscimol and adjacent Nissl stained sections. Bottom plot: the intensity of biotin staining as a function of distance from the midline (1 hemisphere from a tempo bird; 8 hemispheres from 4 pitch birds). The thick black line is from the example sections shown above. L: field L; CM: caudal mesopallium. Scale bar = 1 mm NisslBiotin Kim et al. Molecular Brain (2023) 16:48 Page 4 of 5 ANOVA). We also did not observe significant changes even cell types [18], could reveal relationships between in performance during unilateral infusion of muscimol spatially segregated sound feature encoding in the audi- in both groups (Fig.  1C–F; tempo: p > 0.2, n = 3 birds; tory cortex and the processing of fundamental percepts pitch: p > 0.7, n = 4 birds, Tukey–Kramer post-hoc tests, of complex sounds. following ANOVA with pre-surgery, PBS, unilateral and bilateral muscimol conditions). Abbreviations In this study, we investigated the link between auditory STRF Spectrotemporal receptive fields tuning properties and perception of complex sounds. We PBS Phosphate buffered saline asked whether the lateral subregion of the songbird pri- mary auditory cortex, which exhibits broadband spec- Supplementary Information tral tuning, contributes specifically to tempo processing, The online version contains supplementary material available at https:// doi. org/ 10. 1186/ s13041‑ 023‑ 01039‑5. without affecting pitch discrimination. Our results show that reversible bilateral inactivation of lateral field L Additional file 1. Methods. caused a significant reduction in performance in both Additional file 2: Figure S1. Performance comparison across different tempo and pitch discrimination. Therefore, while prior magnitudes of tempo and pitch shift before implant surgery. (A–B) Prob‑ work has shown that the spectrotemporal receptive fields ability of correct trials for different tempo (A) or pitch shifts (B) (mean ± SEM). Different shades of gray for bars indicate difficulty and correspond of neurons in the songbird auditory cortex are spatially to those in Fig. 1C and D. Performance was not significantly different organized, our results do not provide strong support for across different magnitude of shifts for either group (tempo group: F(5,18) the hypothesis that different subregions of auditory cor - = 2, p = 0.13; pitch group: F (5,18) = 2.64, p = 0.059, ANOVA). Figure S2. Performance of individual birds during bilateral muscimol infusion. (A–B) tex subserve different percepts. Probability of correct trials for different tempo (A) and pitch shifts (B). The Several limitations affect our ability to make strong blue and red lines show the average (± SEM) P(correct) for saline and conclusions about whether different subregions of field bilateral muscimol conditions, respectively (n = 4 birds on tempo task; n = 4 birds on pitch task). The gray lines show P (correct) for individual birds L differentially contribute to basic perceptual qualities during bilateral muscimol infusions, and the dashed lines indicate the per‑ of sound. First, we did not perform the converse experi- formance of the two example birds shown in Fig. 1C and D, respectively. ment—transient inactivation of the medial subregion of Field L—to test whether it differentially affects pitch vs. Acknowledgements tempo processing. Second, our muscimol infusion in We thank Adria Arteseros and Hyesook Lee for technical assistance. the lateral field L spread dorsoventrally across all lay - Author contributions ers of Field L and part of the caudal mesopallium (CM), GK conceived and performed experiments, and analyzed data. MS‑ V imaged a secondary cortical region, affecting neurons that may and analyzed histological data. GK, MS‑ V, and MHK wrote the manuscript. All play different roles in tempo and pitch processing due authors read and approved the final manuscript. to their markedly different temporal tuning properties Funding [7, 14, 15]. Third, we did not perform neural recordings This work was supported by NIH Grant R03DC012428, KBRI basic research during muscimol infusion to confirm whether musci - program through Korea Brain Research Institute funded by Ministry of Science and ICT of Korea (23‑BR‑01‑01, 23‑BR‑04‑04, and 22‑BR‑03‑07), and Basic mol inactivated lateral neurons while sparing the tun- Research Program of the National Research Foundation of Korea funded by ing properties of medial neurons. Although the above the Ministry of Science and ICT of Korea (NRF‑2021R1F1A1049434) to G.K, chance level performance during muscimol infusion Korea Brain Research Institute funded by Ministry of Science and ICT of Korea (23‑BR‑02‑10) to M.S‑ V, NIH grant R01DC011356 to Allison Doupe, and NIH and our histological quantification (Fig.  1G) indicate Grant R01NS129695 to M.H.K. that a substantial medial portion of field L was spared of muscimol, neural recordings would provide more Availability of data and materials All data and materials are available from the corresponding author upon direct examination of the muscimol effect, includ - reasonable request. ing potential changes in tuning via local connections. Finally, while a global gradient in the receptive fields Declarations of neurons in field L has been observed in songbirds, more recent studies in birds and mammals have found Ethics approval and consent to participate All procedures were performed in accordance with protocols approved by local heterogeneity in response properties of auditory the University of California, San Francisco Institutional Animal Care and Use neurons [16, 17], raising the possibility that local het- Committee (AN107972). erogeneity in spectral tuning widths may counteract the Consent for publication effect of inactivation based on a global gradient. Future Not applicable. experiments with a better spatial control of neural activity, such as optogenetic manipulations combined Competing interests The authors declare that they have no competing interests. with genetic targeting of specific regions, layers, or K im et al. Molecular Brain (2023) 16:48 Page 5 of 5 Received: 29 December 2022 Accepted: 25 May 2023 References 1. Eggermont JJ, Johannesma PM, Aertsen AM. Reverse‑ correlation meth‑ ods in auditory research. Q Rev Biophys. 1983;16:341–414. 2. Theunissen FE, Sen K, Doupe AJ. Spectral‑temporal receptive fields of nonlinear auditory neurons obtained using natural sounds. J Neurosci. 2000;20:2315–31. 3. Woolley SMN, Gill PR, Fremouw T, Theunissen FE. Functional groups in the avian auditory system. J Neurosci. 2009;29:2780–93. 4. Moore JM, Woolley SMN. Emergent tuning for learned vocalizations in auditory cortex. Nat Neurosci. 2019;22:1469–76. 5. Nagel KI, Doupe AJ. Organizing principles of spectro‑temporal encoding in the avian primary auditory area field L. Neuron. 2008;58:938–55. 6. Nagel K, Kim G, McLendon H, Doupe A. A bird brain’s view of auditory processing and perception. Hear Res. 2011;273:123–33. 7. Kim G, Doupe A. Organized representation of spectrotemporal features in songbird auditory forebrain. J Neurosci. 2011;31:16977–90. 8. Nagel KI, McLendon HM, Doupe AJ. Differential influence of frequency, timing, and intensity cues in a complex acoustic categorization task. J Neurophysiol. 2010;104:1426–37. 9. Elie JE, Theunissen FE. Zebra finches identify individuals using vocal signatures unique to each call type. Nat Commun. 2018;9:1–11. 10. Woolley SC, Doupe AJ. Social context‑induced song variation affects female behavior and gene expression. PLoS Biol. 2008;6: e62. 11. Paul A, McLendon H, Rally V, Sakata JT, Woolley SC. Behavioral discrimi‑ nation and time‑series phenotyping of birdsong performance. PLoS Comput Biol. 2021;17: e1008820. 12. Moorman S, Ahn J‑R, Kao MH. Plasticity of stereotyped birdsong driven by chronic manipulation of cortical‑basal ganglia activity. Curr Biol. 2021;31:2619‑2632.e4. 13. Tanaka M, Singh Alvarado J, Murugan M, Mooney R. Focal expression of mutant huntingtin in the songbird basal ganglia disrupts cortico‑ basal ganglia networks and vocal sequences. Proc Natl Acad Sci. 2016;113:E1720–7. 14. Lim Y, Lagoy R, Shinn‑ Cunningham BG, Gardner TJ. Transforma‑ tion of temporal sequences in the zebra finch auditory system. Elife. 2016;5:e18205. 15. Calabrese A, Woolley SMN. Coding principles of the canonical cortical microcircuit in the avian brain. Proc Natl Acad Sci. 2015;112:3517–22. 16. Araki M, Bandi MM, Yazaki‑Sugiyama Y. Mind the gap: neural coding of species identity in birdsong prosody. Science. 2016;354:1282–7. 17. Zeng H, Huang J, Chen M, Wen Y, Shen Z, Poo M. Local homogeneity of tonotopic organization in the primary auditory cortex of marmosets. Proc Natl Acad Sci. 2019;116:3239–44. 18. Lee C, Lavoie A, Liu J, Chen SX, Liu B. Light up the brain: the application of optogenetics in cell‑type specific dissection of mouse brain circuits. Front Neural Circuits. 2020. https:// doi. org/ 10. 3389/ fncir. 2020. 00018. Publisher’s Note Springer Nature remains neutral with regard to jurisdictional claims in pub‑ lished maps and institutional affiliations. Re Read ady y to to submit y submit your our re researc search h ? Choose BMC and benefit fr ? Choose BMC and benefit from om: : fast, convenient online submission thorough peer review by experienced researchers in your field rapid publication on acceptance support for research data, including large and complex data types • gold Open Access which fosters wider collaboration and increased citations maximum visibility for your research: over 100M website views per year At BMC, research is always in progress. Learn more biomedcentral.com/submissions

Journal

Molecular Brain – Springer Journals

Published: Jun 3, 2023

Keywords: Auditory perception; Auditory processing; Spectrotemporal receptive fields; Songbirds; Operant behavior

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