Protein kinase A in the pedunculopontine tegmental nucleus of rat contributes to regulation of rapid eye movement sleep - PubMed (original) (raw)

Figure 4.

Relationship between recovery REM sleep and activation of PKA in the PPT. A, Representative agarose gel, visualized with UV light, showing the activity profile of the PKA in the PPT from UR-VC-UR, RD-VC-UR, RD-D1-UR, RD-D2-UR, RD-D3-UR, and RD-VC-RD groups of animals (note that the lanes are random; the technician performing Western blotting and densitometric analysis was blind to experimental group). The bottom band represents phosphorylated fluorescent-labeled Kemptide (P), and the top band represents the remaining unphosphorylated peptide (S). In the gel, phosphorylated product migrated toward the anode (+), and unphosphorylated peptide migrated toward the cathode (−). −Ve Control, no protein or enzyme added; +Ve control, Purified catalytic subunit of PKA (20 ng); in each experimental sample 2 μg of protein added. B, Data from densitometric analysis of PPT PKA activity are expressed as a percentage of control (Gr. 1, UR-VC-UR). Each bar represents PPT PKA activity level (mean ± SE) of the animals (n = 8/group) in group 1 (UR-VC-UR), group 2 (RD-VC-UR), group 3 (RD-D1-UR), group 4 (RD-D2-UR), and group 5 (RD-D3-UR). Note that 2 h after the selective REM sleep deprivation and microinjections of vehicle control, PPT PKA activity levels increased (group 2). Also note that after microinjections of three different concentrations (0.55, 1.1, and 2.2 nmol/100 nl) of Rp-CAMPS into the PPT in three different groups of animals (groups 3, 4, and 5), PPT PKA activity levels decreased in a dose-dependent manner. Post hoc tests (Bonferroni's multiple-comparison test): ***p < 0.001, compared with group 1 (UR-VC-UR); ΔΔΔ_p_ < 0.001, compared with group 2 (RD-VC-UR). C–E, The level of PKA activity in the PPT for each animal (n = 40 rats) is shown as a function of the percentage of time spent in REM sleep (C), wakefulness (D), and SWS (E) during the 2 h period (12:00 P.M. to 2:00 P.M.) of recovery sleep–wake (S–W) cycle. Because some of the data points have identical values, not all 40 data points are visible in C–E. C, Plot of linear regression best fit (solid line; Pearsons product-moment correlation) shows a statistically significant positive slope (Rsqr = 0.944; p < 0.001). These data indicate that the level of PKA activity in the PPT depends positively on the percentage of recovery REM sleep in the period of recovery S–W cycle. D, Plot of linear regression best fit did not reveal any significant relationship between the levels of PKA activity in the PPT and total percentages of time spent in wakefulness (Rsqr = 0.125; p = 0.373). These data indicate that the level of PKA activity in the PPT did not vary as a function of wakefulness in the period of recovery S–W cycle. E, Plot of linear regression best fit revealed a significant negative relationship between levels of PKA activity in the PPT and total percentages of time spent in SWS (Rsqr = 0.903; p < 0.001). These data indicate that the level of PKA activity in the PPT depends negatively on the percentage of SWS in the period of recovery S–W cycle. UR-VC-UR, unrestricted REM sleep-vehicle control-unrestricted REM sleep; RD-VC-UR, REM sleep deprivation-vehicle control-unrestricted REM sleep; RD-D1-UR, REM sleep deprivation-dose 1 Rp-CAMPS-unrestricted REM sleep; RD-D2-UR, REM sleep deprivation-dose 2 Rp-CAMPS-unrestricted REM sleep; RD-D3-UR, REM sleep deprivation-dose 3 Rp-CAMPS-unrestricted REM sleep; RD-VC-RD, REM sleep deprivation-vehicle control-REM sleep deprivation.