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Coexpression analysis of nine neuropeptides in the neurosecretory preoptic area of larval zebrafish.

Herget U, Ryu S - Front Neuroanat (2015)

Bottom Line: To identify distinct cell types present in the larval NPO, we also generated a comprehensive 3D map of 9 zebrafish homologs of typical neuropeptides found in the mammalian PVN (arginine vasopressin (AVP), corticotropin-releasing hormone (CRH), proenkephalin a (penka)/b (penkb), neurotensin (NTS), oxytocin (OXT), vasoactive intestinal peptide (VIP), cholecystokinin (CCK), and somatostatin (SST)).Our results allowed the subclassification of NPO cell types, and differences in variability of coexpression profiles suggest potential targets of biochemical plasticity.Thus, this work provides an important basis for the analysis of the development, function, and plasticity of the primary neuroendocrine brain region in larval zebrafish.

View Article: PubMed Central - PubMed

Affiliation: Developmental Genetics of the Nervous System, Max Planck Institute for Medical Research Heidelberg, Germany ; The Hartmut Hoffmann-Berling International Graduate School of Molecular and Cellular Biology, University of Heidelberg Heidelberg, Germany.

ABSTRACT
The paraventricular nucleus (PVN) of the hypothalamus in mammals coordinates neuroendocrine, autonomic and behavioral responses pivotal for homeostasis and the stress response. A large amount of studies in rodents has documented that the PVN contains diverse neuronal cell types which can be identified by the expression of distinct secretory neuropeptides. Interestingly, PVN cell types often coexpress multiple neuropeptides whose relative coexpression levels are subject to environment-induced plasticity. Due to their small size and transparency, zebrafish larvae offer the possibility to comprehensively study the development and plasticity of the PVN in large groups of intact animals, yet important anatomical information about the larval zebrafish PVN-homologous region has been missing. Therefore we recently defined the location and borders of the larval neurosecretory preoptic area (NPO) as the PVN-homologous region in larval zebrafish based on transcription factor expression and cell type clustering. To identify distinct cell types present in the larval NPO, we also generated a comprehensive 3D map of 9 zebrafish homologs of typical neuropeptides found in the mammalian PVN (arginine vasopressin (AVP), corticotropin-releasing hormone (CRH), proenkephalin a (penka)/b (penkb), neurotensin (NTS), oxytocin (OXT), vasoactive intestinal peptide (VIP), cholecystokinin (CCK), and somatostatin (SST)). Here we extend this chemoarchitectural map to include the degrees of coexpression of two neuropeptides in the same cell by performing systematic pairwise comparisons. Our results allowed the subclassification of NPO cell types, and differences in variability of coexpression profiles suggest potential targets of biochemical plasticity. Thus, this work provides an important basis for the analysis of the development, function, and plasticity of the primary neuroendocrine brain region in larval zebrafish.

No MeSH data available.


Cell type staining combinations showing no coexpression. (A) Cells expressing avp(A’) or vip(A”) form neighboring but separate clusters. (B) Cells expressing crh(B’) or vip(B”) are similarly separated. (C) Cells expressing cck cluster rostrally (C’), and are therefore distant from vip-positive cells (C”). (D) Cells expressing sst1.1(D’) also cluster in a separate region from vip-positive cells (D”). (E)cck-positive cells (E’) are rostrally neighboring crh-positive cells (E”). (F) Cells expressing cck(F’) or nts(F”) are separate. (G)penka-positive cells (G’) are scattered, but do not overlap with vip expression (G”). (H)oxt-positive (H’) and cck-positive cells (H”) are also separated. (I) The clusters formed by cells expressing oxt(I’) or vip(I”) do not show coexpression. (J)vip(J’) and penkb(J”) are also not coexpressed. (K) The rostral location of cck-positive cells (K’) is separate from the neighboring avp-positive cluster (K”). (L)penka-positive cells (L’) also are close to the cck-positive population (L”), but still separate. (M) Cells positive for penkb(M’) surround the central nts-positive cluster (M”). (N) Cells expressing avp(N’) or oxt(N”) are intermingled, but the peptides are not coexpressed (single planes: O–O”). (P) Cells expressing nts(P’) or oxt(P”) have similar locations, but these peptides are not coexpressed. (Q) Cells expressing oxt(Q’) or sst1.1(Q”) are also intermingled and do not coexpress these peptides (single planes: R–R”). All images show maximum intensity projections, unless indicated otherwise. Scale bar: 50 μm.
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Figure 2: Cell type staining combinations showing no coexpression. (A) Cells expressing avp(A’) or vip(A”) form neighboring but separate clusters. (B) Cells expressing crh(B’) or vip(B”) are similarly separated. (C) Cells expressing cck cluster rostrally (C’), and are therefore distant from vip-positive cells (C”). (D) Cells expressing sst1.1(D’) also cluster in a separate region from vip-positive cells (D”). (E)cck-positive cells (E’) are rostrally neighboring crh-positive cells (E”). (F) Cells expressing cck(F’) or nts(F”) are separate. (G)penka-positive cells (G’) are scattered, but do not overlap with vip expression (G”). (H)oxt-positive (H’) and cck-positive cells (H”) are also separated. (I) The clusters formed by cells expressing oxt(I’) or vip(I”) do not show coexpression. (J)vip(J’) and penkb(J”) are also not coexpressed. (K) The rostral location of cck-positive cells (K’) is separate from the neighboring avp-positive cluster (K”). (L)penka-positive cells (L’) also are close to the cck-positive population (L”), but still separate. (M) Cells positive for penkb(M’) surround the central nts-positive cluster (M”). (N) Cells expressing avp(N’) or oxt(N”) are intermingled, but the peptides are not coexpressed (single planes: O–O”). (P) Cells expressing nts(P’) or oxt(P”) have similar locations, but these peptides are not coexpressed. (Q) Cells expressing oxt(Q’) or sst1.1(Q”) are also intermingled and do not coexpress these peptides (single planes: R–R”). All images show maximum intensity projections, unless indicated otherwise. Scale bar: 50 μm.

Mentions: Many of the neuropeptide combinations (16/36) showed no coexpression in the same cell, and often these neuropeptides were expressed in spatially separate clusters. The rostralmost clusters formed by cells expressing avp, crh, cck, or sst1.1 did not show any overlap with the caudalmost cluster formed by cells expressing vip (Figures 2A–D”, 5–8 animals analyzed). Among the rostral group, crh and cck were not coexpressed in the same cells, and cck expression was also separate from the large nts-positive cluster (Figures 2E–F”, 6–7 animals analyzed). The penka-expressing cluster extended as far caudally as the small group of vip-positive cells, which however occupied a more lateral region (Figures 2G–G”, 8 animals analyzed). oxt-expressing cells formed a large central cluster, and they did not overlap with the rostrally located cck-expressing cells, nor with the caudally located vip-expressing group (Figures 2H–I”, 6–9 animals analyzed). The vip-positive cluster was also separate from cells expressing penkb (Figures 2J–J”, 7 animals analyzed). The rostral cluster of cck-positive cells was close to, but separate from the avp-positive and penka-positive clusters (Figures 2K–L”, 7 animals analyzed). penkb-positive cells surrounded the more central nts-positive cluster (Figures 2M–M”, 5 animals analyzed). Within the center of the NPO, cells expressing avp or oxt were found intermingled in the same region, but did not overlap (Figures 2N–O”, 8 animals analyzed). The central and intermingled clusters of cells expressing oxt or nts did not show coexpression of these two peptides (Figures 2P–P”, 10 animals analyzed). Similar spatial proximity without coexpression was also found for cells expressing oxt or sst1.1 (Figures 2Q–R”, 8 animals analyzed).


Coexpression analysis of nine neuropeptides in the neurosecretory preoptic area of larval zebrafish.

Herget U, Ryu S - Front Neuroanat (2015)

Cell type staining combinations showing no coexpression. (A) Cells expressing avp(A’) or vip(A”) form neighboring but separate clusters. (B) Cells expressing crh(B’) or vip(B”) are similarly separated. (C) Cells expressing cck cluster rostrally (C’), and are therefore distant from vip-positive cells (C”). (D) Cells expressing sst1.1(D’) also cluster in a separate region from vip-positive cells (D”). (E)cck-positive cells (E’) are rostrally neighboring crh-positive cells (E”). (F) Cells expressing cck(F’) or nts(F”) are separate. (G)penka-positive cells (G’) are scattered, but do not overlap with vip expression (G”). (H)oxt-positive (H’) and cck-positive cells (H”) are also separated. (I) The clusters formed by cells expressing oxt(I’) or vip(I”) do not show coexpression. (J)vip(J’) and penkb(J”) are also not coexpressed. (K) The rostral location of cck-positive cells (K’) is separate from the neighboring avp-positive cluster (K”). (L)penka-positive cells (L’) also are close to the cck-positive population (L”), but still separate. (M) Cells positive for penkb(M’) surround the central nts-positive cluster (M”). (N) Cells expressing avp(N’) or oxt(N”) are intermingled, but the peptides are not coexpressed (single planes: O–O”). (P) Cells expressing nts(P’) or oxt(P”) have similar locations, but these peptides are not coexpressed. (Q) Cells expressing oxt(Q’) or sst1.1(Q”) are also intermingled and do not coexpress these peptides (single planes: R–R”). All images show maximum intensity projections, unless indicated otherwise. Scale bar: 50 μm.
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Figure 2: Cell type staining combinations showing no coexpression. (A) Cells expressing avp(A’) or vip(A”) form neighboring but separate clusters. (B) Cells expressing crh(B’) or vip(B”) are similarly separated. (C) Cells expressing cck cluster rostrally (C’), and are therefore distant from vip-positive cells (C”). (D) Cells expressing sst1.1(D’) also cluster in a separate region from vip-positive cells (D”). (E)cck-positive cells (E’) are rostrally neighboring crh-positive cells (E”). (F) Cells expressing cck(F’) or nts(F”) are separate. (G)penka-positive cells (G’) are scattered, but do not overlap with vip expression (G”). (H)oxt-positive (H’) and cck-positive cells (H”) are also separated. (I) The clusters formed by cells expressing oxt(I’) or vip(I”) do not show coexpression. (J)vip(J’) and penkb(J”) are also not coexpressed. (K) The rostral location of cck-positive cells (K’) is separate from the neighboring avp-positive cluster (K”). (L)penka-positive cells (L’) also are close to the cck-positive population (L”), but still separate. (M) Cells positive for penkb(M’) surround the central nts-positive cluster (M”). (N) Cells expressing avp(N’) or oxt(N”) are intermingled, but the peptides are not coexpressed (single planes: O–O”). (P) Cells expressing nts(P’) or oxt(P”) have similar locations, but these peptides are not coexpressed. (Q) Cells expressing oxt(Q’) or sst1.1(Q”) are also intermingled and do not coexpress these peptides (single planes: R–R”). All images show maximum intensity projections, unless indicated otherwise. Scale bar: 50 μm.
Mentions: Many of the neuropeptide combinations (16/36) showed no coexpression in the same cell, and often these neuropeptides were expressed in spatially separate clusters. The rostralmost clusters formed by cells expressing avp, crh, cck, or sst1.1 did not show any overlap with the caudalmost cluster formed by cells expressing vip (Figures 2A–D”, 5–8 animals analyzed). Among the rostral group, crh and cck were not coexpressed in the same cells, and cck expression was also separate from the large nts-positive cluster (Figures 2E–F”, 6–7 animals analyzed). The penka-expressing cluster extended as far caudally as the small group of vip-positive cells, which however occupied a more lateral region (Figures 2G–G”, 8 animals analyzed). oxt-expressing cells formed a large central cluster, and they did not overlap with the rostrally located cck-expressing cells, nor with the caudally located vip-expressing group (Figures 2H–I”, 6–9 animals analyzed). The vip-positive cluster was also separate from cells expressing penkb (Figures 2J–J”, 7 animals analyzed). The rostral cluster of cck-positive cells was close to, but separate from the avp-positive and penka-positive clusters (Figures 2K–L”, 7 animals analyzed). penkb-positive cells surrounded the more central nts-positive cluster (Figures 2M–M”, 5 animals analyzed). Within the center of the NPO, cells expressing avp or oxt were found intermingled in the same region, but did not overlap (Figures 2N–O”, 8 animals analyzed). The central and intermingled clusters of cells expressing oxt or nts did not show coexpression of these two peptides (Figures 2P–P”, 10 animals analyzed). Similar spatial proximity without coexpression was also found for cells expressing oxt or sst1.1 (Figures 2Q–R”, 8 animals analyzed).

Bottom Line: To identify distinct cell types present in the larval NPO, we also generated a comprehensive 3D map of 9 zebrafish homologs of typical neuropeptides found in the mammalian PVN (arginine vasopressin (AVP), corticotropin-releasing hormone (CRH), proenkephalin a (penka)/b (penkb), neurotensin (NTS), oxytocin (OXT), vasoactive intestinal peptide (VIP), cholecystokinin (CCK), and somatostatin (SST)).Our results allowed the subclassification of NPO cell types, and differences in variability of coexpression profiles suggest potential targets of biochemical plasticity.Thus, this work provides an important basis for the analysis of the development, function, and plasticity of the primary neuroendocrine brain region in larval zebrafish.

View Article: PubMed Central - PubMed

Affiliation: Developmental Genetics of the Nervous System, Max Planck Institute for Medical Research Heidelberg, Germany ; The Hartmut Hoffmann-Berling International Graduate School of Molecular and Cellular Biology, University of Heidelberg Heidelberg, Germany.

ABSTRACT
The paraventricular nucleus (PVN) of the hypothalamus in mammals coordinates neuroendocrine, autonomic and behavioral responses pivotal for homeostasis and the stress response. A large amount of studies in rodents has documented that the PVN contains diverse neuronal cell types which can be identified by the expression of distinct secretory neuropeptides. Interestingly, PVN cell types often coexpress multiple neuropeptides whose relative coexpression levels are subject to environment-induced plasticity. Due to their small size and transparency, zebrafish larvae offer the possibility to comprehensively study the development and plasticity of the PVN in large groups of intact animals, yet important anatomical information about the larval zebrafish PVN-homologous region has been missing. Therefore we recently defined the location and borders of the larval neurosecretory preoptic area (NPO) as the PVN-homologous region in larval zebrafish based on transcription factor expression and cell type clustering. To identify distinct cell types present in the larval NPO, we also generated a comprehensive 3D map of 9 zebrafish homologs of typical neuropeptides found in the mammalian PVN (arginine vasopressin (AVP), corticotropin-releasing hormone (CRH), proenkephalin a (penka)/b (penkb), neurotensin (NTS), oxytocin (OXT), vasoactive intestinal peptide (VIP), cholecystokinin (CCK), and somatostatin (SST)). Here we extend this chemoarchitectural map to include the degrees of coexpression of two neuropeptides in the same cell by performing systematic pairwise comparisons. Our results allowed the subclassification of NPO cell types, and differences in variability of coexpression profiles suggest potential targets of biochemical plasticity. Thus, this work provides an important basis for the analysis of the development, function, and plasticity of the primary neuroendocrine brain region in larval zebrafish.

No MeSH data available.