Transiently increasing cAMP levels selectively in hippocampal excitatory neurons during sleep deprivation prevents memory deficits caused by sleep loss

Abstract

The hippocampus is particularly sensitive to sleep loss. Although previous work has indicated that sleep deprivation impairs hippocampal cAMP signaling, it remains to be determined whether the cognitive deficits associated with sleep deprivation are caused by attenuated cAMP signaling in the hippocampus. Further, it is unclear which cell types are responsible for the memory impairments associated with sleep deprivation. Transgenic approaches lack the spatial resolution to manipulate specific signaling pathways selectively in the hippocampus, while pharmacological strategies are limited in terms of cell-type specificity. Therefore, we used a pharmacogenetic approach based on a virus-mediated expression of a Gαs-coupled Drosophila octopamine receptor selectively in mouse hippocampal excitatory neurons in vivo. With this approach, a systemic injection with the receptor ligand octopamine leads to increased cAMP levels in this specific set of hippocampal neurons. We assessed whether transiently increasing cAMP levels during sleep deprivation prevents memory consolidation deficits associated with sleep loss in an object-location task. Five hours of total sleep deprivation directly following training impaired the formation of object-location memories. Transiently increasing cAMP levels in hippocampal neurons during the course of sleep deprivation prevented these memory consolidation deficits. These findings demonstrate that attenuated cAMP signaling in hippocampal excitatory neurons is a critical component underlying the memory deficits in hippocampus-dependent learning tasks associated with sleep deprivation.

Keywords: hippocampus; learning; memory; pharmacogenetics; sleep; sleep disruption.

Figure 1.

Viral expression of DmOctβR1 is restricted to excitatory neurons of the hippocampus. A, Mice were injected with pAAV₉-CaMKIIα0.4-EGFP or pAAV₉-CaMKIIα0.4-DmOctβ1R-HA to drive expression of EGFP or the Gαs-coupled DmOctβ1R in hippocampal excitatory neurons. B, Activation of the DmOctβ1R by its ligand octopamine (OCT) stimulates adenylyl cyclase activity, which synthesizes cAMP. C, D, Representative low-magnification images of EGFP and DmOctβ1R expression in coronal brain sections. Scale bar, 570 μm. E, F, Representative images EGFP and DmOctβ1R expression in all three major hippocampal subregions. Scale bar, 200 μm. G–L, Higher-magnification images of area CA1 (E, H), CA3 (F, I), and dentate gyrus (G, J). Scale bar, 20 μm. AC, Adenyl cyclase; ITR, inverted terminal repeats.

Figure 2.

Activation of the DmOctβ1R in hippocampal neurons leads to increased cAMP levels selectively in the hippocampus. A, A systemic injection with octopamine (1 mg/kg) increased hippocampal cAMP levels 30 min after injection (*p = 0.034, t test) in mice expressing DmOctβ1R. B, Systemic injection with octopamine (1 mg/kg) did not alter cAMP levels in the prefrontal cortex 30 min after injection in mice expressing DmOctβ1R (p = 0.668, t test). All data are expressed as the mean ± SEM.

Figure 3.

Increasing neuronal cAMP levels in hippocampal neurons prevents memory deficits caused by sleep deprivation. A, A diagram illustrating the training schedule and time points at which all animals received intraperitonal injections with octopamine. B, Viral expression of the Gαs-coupled DmOctβ1R in hippocampal neurons does not affect object exploration levels during training in the object–place recognition task (effect of virus: F_(1,35) = 1.640, p = 0.209). All groups showed decreased total object exploration time during consecutive training sessions in a similar fashion (effect of session: F_(2,70) = 58.723, p = 0.0001; interaction effect: F_(2,70) = 0.936, p = 0.397). C, Mice expressing EGFP or DmOctβ1R were trained in the hippocampus-dependent object–place recognition task and sleep deprived for 5 h immediately after training [sleep-deprived (SD) animals] or were left undisturbed [non-sleep-deprived (NSD) animals]. Mice received systemic injections with octopamine directly after training and 2.5 h into sleep deprivation. Boosting cAMP signaling by activating the of DmOctβ1R prevents memory deficits caused by sleep deprivation (n = 9–10; effect of sleep deprivation: F_(1,35) = 6.603, p = 0.015; effect of virus: F_(1,35) = 20.256, p < 0.001; interaction effect: F_(1,35) = 4.565, p = 0.04; EGFP SD group vs other groups, p < 0.05). Dotted line indicates chance performance. Error bars denote SEM. *p < 0.05. All data are expressed as the mean ± SEM.