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Neuronal ensembles sufficient for recovery sleep and the sedative actions of α2 adrenergic agonists.

Zhang Z, Ferretti V, Güntan İ, Moro A, Steinberg EA, Ye Z, Zecharia AY, Yu X, Vyssotski AL, Brickley SG, Yustos R, Pillidge ZE, Harding EC, Wisden W, Franks NP - Nat. Neurosci. (2015)

Bottom Line: For the α2 adrenergic receptor agonist dexmedetomidine, we found that sedation and LORR were in fact distinct states, requiring different brain areas: the preoptic hypothalamic area and locus coeruleus (LC), respectively.Instead, we found that dexmedetomidine-induced sedation resembled the deep recovery sleep that follows sleep deprivation.Thus, α2 adrenergic receptor-induced sedation and recovery sleep share hypothalamic circuitry sufficient for producing these behavioral states.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Sciences, Imperial College London, South Kensington, UK.

ABSTRACT
Do sedatives engage natural sleep pathways? It is usually assumed that anesthetic-induced sedation and loss of righting reflex (LORR) arise by influencing the same circuitry to lesser or greater extents. For the α2 adrenergic receptor agonist dexmedetomidine, we found that sedation and LORR were in fact distinct states, requiring different brain areas: the preoptic hypothalamic area and locus coeruleus (LC), respectively. Selective knockdown of α2A adrenergic receptors from the LC abolished dexmedetomidine-induced LORR, but not sedation. Instead, we found that dexmedetomidine-induced sedation resembled the deep recovery sleep that follows sleep deprivation. We used TetTag pharmacogenetics in mice to functionally mark neurons activated in the preoptic hypothalamus during dexmedetomidine-induced sedation or recovery sleep. The neuronal ensembles could then be selectively reactivated. In both cases, non-rapid eye movement sleep, with the accompanying drop in body temperature, was recapitulated. Thus, α2 adrenergic receptor-induced sedation and recovery sleep share hypothalamic circuitry sufficient for producing these behavioral states.

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Dexmedetomidine-induced sedation and recovery sleep induced cFOS expression in overlapping regions of the mouse hypothalamic preoptic area and septum. (a) Schematic of the relevant preoptic hypothalamic and septal areas: left-hand drawing, midline-sagittal section, red line marks position of the section; middle drawing, coronal section, boxed area, magnified on the right, shows the relevant anatomical sites. (b) Line drawings of cFOS protein expression in the boxed area at 30 minutes after saline injection and 30 or 60 minutes after dexmedetomidine (100 μg/kg) injections or 2 hours into recovery sleep after sleep deprivation; black dots represent cFOS-positive cells (see Supplementary Fig. 1 for representative photographs); relative to its expression after a saline injection, the endogenous cfos gene is induced widely in the area by sedative doses of dexmedetomidine or during recovery sleep. (c) Number of cFOS positive neurons in selected anatomical sites after saline (white) or dexmedetomidine (red) injections or recovery sleep (gray). The boxes represent the s.e.m, and the bars show the range of the data. Asterisks represent significance relative to saline *P<0.05, **P<0.01, ***P<0.001 (t-test). LPO, lateral preoptic area; LSV, lateral septum, ventral; MPO, medial preoptic area SHy, septo-hypothalamic nucleus; STLD, stria terminalis lateral dorsal; STMA, stria terminalis medial anterior; VLPO, ventral lateral preoptic area.
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Figure 2: Dexmedetomidine-induced sedation and recovery sleep induced cFOS expression in overlapping regions of the mouse hypothalamic preoptic area and septum. (a) Schematic of the relevant preoptic hypothalamic and septal areas: left-hand drawing, midline-sagittal section, red line marks position of the section; middle drawing, coronal section, boxed area, magnified on the right, shows the relevant anatomical sites. (b) Line drawings of cFOS protein expression in the boxed area at 30 minutes after saline injection and 30 or 60 minutes after dexmedetomidine (100 μg/kg) injections or 2 hours into recovery sleep after sleep deprivation; black dots represent cFOS-positive cells (see Supplementary Fig. 1 for representative photographs); relative to its expression after a saline injection, the endogenous cfos gene is induced widely in the area by sedative doses of dexmedetomidine or during recovery sleep. (c) Number of cFOS positive neurons in selected anatomical sites after saline (white) or dexmedetomidine (red) injections or recovery sleep (gray). The boxes represent the s.e.m, and the bars show the range of the data. Asterisks represent significance relative to saline *P<0.05, **P<0.01, ***P<0.001 (t-test). LPO, lateral preoptic area; LSV, lateral septum, ventral; MPO, medial preoptic area SHy, septo-hypothalamic nucleus; STLD, stria terminalis lateral dorsal; STMA, stria terminalis medial anterior; VLPO, ventral lateral preoptic area.

Mentions: If not at the LC, where in the mouse brain is the α2 receptor-induced sedative response taking place, and could natural sleep pathways be involved? Previous work emphasized that the ventro lateral preoptic (VLPO) nucleus in the PO hypothalamus was activated following dexmedetomidine administration8, and during normal and recovery sleep21,31, so we reviewed cFOS activation in the PO area. Mice were injected with a sedative dose of dexmedetomidine (100 μg/kg) or control saline. Brains were taken either 30 or 60 minutes afterwards, and analyzed for endogenous cFOS expression (Fig. 2 and Supplementary Fig. 1). We also investigated cFOS expression 2 hours into the recovery sleep following 4 hours of sleep deprivation (Fig. 2 and Supplementary Fig. 1). Although for both dexmedetomidine-induced sedation and during recovery sleep, there were some cFOS positive neurons in VLPO, many more activated cells were found in the wider PO area (LPO, MPO) and in a cluster of areas just dorsal of the preoptic region, in the BST (e.g. in the STMA and STLD), and in several septal nuclei: the ventral lateral septum (LSV) and in the septo-hypothalamic nucleus (SHy) (Fig. 2a,b,c). Thus, the broadly similar patterns of induced cFOS during dexmedetomidine-induced sedation and during recovery sleep, indicated that much of the PO region was a suitable location for TetTag-DREADD mapping24-27 to see if activating neurons in this location was sufficient for these behavioral states. We aimed to express, by TetTagging24, a cfos-promoter-inducible hM3Dq-mCHERRY receptor gene25,27 selectively in the PO area of the hypothalamus. The excitatory hM3Dq metabotropic receptor is uniquely activated by the ligand CNO27. By combining this receptor with cfos-dependent TetTagging, the gene encoding the hM3Dq receptor is only turned on following neural activity, and so records, or “tags” ensembles of neurons that have been activated in vivo by a stimulus25. The neurons can then be reactivated later by systemic CNO, allowing sufficiency for the particular behavior25, sleep in this case, to be tested.


Neuronal ensembles sufficient for recovery sleep and the sedative actions of α2 adrenergic agonists.

Zhang Z, Ferretti V, Güntan İ, Moro A, Steinberg EA, Ye Z, Zecharia AY, Yu X, Vyssotski AL, Brickley SG, Yustos R, Pillidge ZE, Harding EC, Wisden W, Franks NP - Nat. Neurosci. (2015)

Dexmedetomidine-induced sedation and recovery sleep induced cFOS expression in overlapping regions of the mouse hypothalamic preoptic area and septum. (a) Schematic of the relevant preoptic hypothalamic and septal areas: left-hand drawing, midline-sagittal section, red line marks position of the section; middle drawing, coronal section, boxed area, magnified on the right, shows the relevant anatomical sites. (b) Line drawings of cFOS protein expression in the boxed area at 30 minutes after saline injection and 30 or 60 minutes after dexmedetomidine (100 μg/kg) injections or 2 hours into recovery sleep after sleep deprivation; black dots represent cFOS-positive cells (see Supplementary Fig. 1 for representative photographs); relative to its expression after a saline injection, the endogenous cfos gene is induced widely in the area by sedative doses of dexmedetomidine or during recovery sleep. (c) Number of cFOS positive neurons in selected anatomical sites after saline (white) or dexmedetomidine (red) injections or recovery sleep (gray). The boxes represent the s.e.m, and the bars show the range of the data. Asterisks represent significance relative to saline *P<0.05, **P<0.01, ***P<0.001 (t-test). LPO, lateral preoptic area; LSV, lateral septum, ventral; MPO, medial preoptic area SHy, septo-hypothalamic nucleus; STLD, stria terminalis lateral dorsal; STMA, stria terminalis medial anterior; VLPO, ventral lateral preoptic area.
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Figure 2: Dexmedetomidine-induced sedation and recovery sleep induced cFOS expression in overlapping regions of the mouse hypothalamic preoptic area and septum. (a) Schematic of the relevant preoptic hypothalamic and septal areas: left-hand drawing, midline-sagittal section, red line marks position of the section; middle drawing, coronal section, boxed area, magnified on the right, shows the relevant anatomical sites. (b) Line drawings of cFOS protein expression in the boxed area at 30 minutes after saline injection and 30 or 60 minutes after dexmedetomidine (100 μg/kg) injections or 2 hours into recovery sleep after sleep deprivation; black dots represent cFOS-positive cells (see Supplementary Fig. 1 for representative photographs); relative to its expression after a saline injection, the endogenous cfos gene is induced widely in the area by sedative doses of dexmedetomidine or during recovery sleep. (c) Number of cFOS positive neurons in selected anatomical sites after saline (white) or dexmedetomidine (red) injections or recovery sleep (gray). The boxes represent the s.e.m, and the bars show the range of the data. Asterisks represent significance relative to saline *P<0.05, **P<0.01, ***P<0.001 (t-test). LPO, lateral preoptic area; LSV, lateral septum, ventral; MPO, medial preoptic area SHy, septo-hypothalamic nucleus; STLD, stria terminalis lateral dorsal; STMA, stria terminalis medial anterior; VLPO, ventral lateral preoptic area.
Mentions: If not at the LC, where in the mouse brain is the α2 receptor-induced sedative response taking place, and could natural sleep pathways be involved? Previous work emphasized that the ventro lateral preoptic (VLPO) nucleus in the PO hypothalamus was activated following dexmedetomidine administration8, and during normal and recovery sleep21,31, so we reviewed cFOS activation in the PO area. Mice were injected with a sedative dose of dexmedetomidine (100 μg/kg) or control saline. Brains were taken either 30 or 60 minutes afterwards, and analyzed for endogenous cFOS expression (Fig. 2 and Supplementary Fig. 1). We also investigated cFOS expression 2 hours into the recovery sleep following 4 hours of sleep deprivation (Fig. 2 and Supplementary Fig. 1). Although for both dexmedetomidine-induced sedation and during recovery sleep, there were some cFOS positive neurons in VLPO, many more activated cells were found in the wider PO area (LPO, MPO) and in a cluster of areas just dorsal of the preoptic region, in the BST (e.g. in the STMA and STLD), and in several septal nuclei: the ventral lateral septum (LSV) and in the septo-hypothalamic nucleus (SHy) (Fig. 2a,b,c). Thus, the broadly similar patterns of induced cFOS during dexmedetomidine-induced sedation and during recovery sleep, indicated that much of the PO region was a suitable location for TetTag-DREADD mapping24-27 to see if activating neurons in this location was sufficient for these behavioral states. We aimed to express, by TetTagging24, a cfos-promoter-inducible hM3Dq-mCHERRY receptor gene25,27 selectively in the PO area of the hypothalamus. The excitatory hM3Dq metabotropic receptor is uniquely activated by the ligand CNO27. By combining this receptor with cfos-dependent TetTagging, the gene encoding the hM3Dq receptor is only turned on following neural activity, and so records, or “tags” ensembles of neurons that have been activated in vivo by a stimulus25. The neurons can then be reactivated later by systemic CNO, allowing sufficiency for the particular behavior25, sleep in this case, to be tested.

Bottom Line: For the α2 adrenergic receptor agonist dexmedetomidine, we found that sedation and LORR were in fact distinct states, requiring different brain areas: the preoptic hypothalamic area and locus coeruleus (LC), respectively.Instead, we found that dexmedetomidine-induced sedation resembled the deep recovery sleep that follows sleep deprivation.Thus, α2 adrenergic receptor-induced sedation and recovery sleep share hypothalamic circuitry sufficient for producing these behavioral states.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Sciences, Imperial College London, South Kensington, UK.

ABSTRACT
Do sedatives engage natural sleep pathways? It is usually assumed that anesthetic-induced sedation and loss of righting reflex (LORR) arise by influencing the same circuitry to lesser or greater extents. For the α2 adrenergic receptor agonist dexmedetomidine, we found that sedation and LORR were in fact distinct states, requiring different brain areas: the preoptic hypothalamic area and locus coeruleus (LC), respectively. Selective knockdown of α2A adrenergic receptors from the LC abolished dexmedetomidine-induced LORR, but not sedation. Instead, we found that dexmedetomidine-induced sedation resembled the deep recovery sleep that follows sleep deprivation. We used TetTag pharmacogenetics in mice to functionally mark neurons activated in the preoptic hypothalamus during dexmedetomidine-induced sedation or recovery sleep. The neuronal ensembles could then be selectively reactivated. In both cases, non-rapid eye movement sleep, with the accompanying drop in body temperature, was recapitulated. Thus, α2 adrenergic receptor-induced sedation and recovery sleep share hypothalamic circuitry sufficient for producing these behavioral states.

Show MeSH
Related in: MedlinePlus