Our research team successfully implanted a sophisticated quadrupole electrode into a monkey, specifically targeting the Ox/Hcrt neurons located in the lateral hypothalamus area (LHA). Confirmation of this precise placement was achieved through meticulous postmortem immunohistology, as illustrated in Figures 1a and 1b. The implanted electrode was connected to a subcutaneous device that facilitated both continuous high-frequency stimulation (HFS) at 80 Hz and low-frequency stimulation (LFS) at 20 Hz, enabling a comprehensive study of neural activity. To induce a parkinsonian state in the monkey, we administered 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) consistently over a period, resulting in stable motor symptoms characterized by a parkinsonian score of 14.6 ± 0.2. These symptoms included bradykinesia, postural instability, and a notable decrease in general activity, which persisted for a duration of 8 consecutive weeks, as detailed in the methods section. Postmortem immunohistology corroborated significant dopaminergic depletion, displaying a dramatic reduction in tyrosine hydroxylase expression within both the striatum and substantia nigra, depicted in Figure 1c.
To assess the sleepiness level, we conducted a minimum of five modified multiple sleep latency tests (mMSLT) on the monkey. Each test comprised three consecutive 20-minute light-OFF sessions scheduled for the morning hours at 10:00, 11:00, and 12:00, with light turned ON briefly between sessions to ensure appropriate conditions. The animal was acclimated in the behavioral cage starting at 9:20 to minimize stress and enhance accuracy. The mMSLT protocol was administered under three conditions: stim-OFF, LFS-ON, and HFS-ON, while also comparing results from healthy and parkinsonian states, as shown in Figure 1d. The electrodes, integrated with a radiotelemetry transmitter, enabled the recording of critical EEG, EOG, and EMG signals under free-moving conditions, essential for accurately detecting different stages of wakefulness and sleep. During the mMSLT protocol, we notably observed the wake and sleep stages restricted to active wake, quiet wake, and light sleep stages 1 and 2, which are detailed in Figure 1e.
Fig. 1
The surgical report and parkinsonian model are as follows: (a), The X-ray images confirm the final placement of the electrode in the LHA, presented in both coronal and sagittal views alongside key internal landmarks characterized by the anterior-posterior commissure (AC-PC) line established through ventriculography (notably, LV indicates the lateral ventricle). (b), Cryostat photographs illustrate a coronal plane representation showing the trajectory and reconstruction of the electrode within the lateral hypothalamic area, precisely -3 mm from the AC line, targeting OxA/Hcrt1 neurons. The enclosed image highlights the crucial immunostaining of OxA/Hcrt1 neurons, with a scale bar measuring 100 μm. (c), Tyrosine hydroxylase (TH) immunostaining is clearly depicted at the striatum (scale bar = 2000 μm) and substantia nigra (scale bar = 1000 μm), contrasting a control animal with an MPTP-treated animal. This section also includes a graph illustrating the percentage of loss of TH staining observed in the caudate (Cd), putamen (Pu), and substantia nigra compacta (SNc) compared to control specimens. d, The design of the modified multiple sleep latency test (mMSLT) shows the structured protocol involving 3 consecutive 20-minute light-OFF sessions, detailed for the times of 10:00 [1], 11:00 [2], and 12:00 [3], conducted in the behavioral cage under varying conditions: stim-OFF or continuous LHA-DBS; LFS-ON (20 Hz) or HFS-ON (80 Hz). e, Example epochs of 30 seconds showcase the EEG, EOG, and EMG signals collected during wake and sleep stages investigated in this study; wake states comprise active and quiet wake, while sleep states include light sleep stages 1 and 2. Important to note are the key anatomical references AC-PC line, LV: lateral ventricle, Pu: putamen, Cd: caudate, and SNc: substantia nigra compacta.
In the initial healthy state, the monkey exhibited spontaneous sleep during 20% of the light-OFF sessions when under the stim-OFF condition; intriguingly, while the occurrence of at least one sleep episode per session exhibited no significant difference in the LFS condition (25%), it drastically increased to 83% under the HFS condition, as indicated in Figure 2a. In the stim-OFF condition, these sleep episodes presented a mean latency of about 19.6 ± 0.2 minutes when all light-OFF sessions were pooled together. This latency was consistently observed across the three successive light-OFF sessions. Under the LFS condition, sleep latency remained statistically comparable to the stim-OFF condition. However, a significant decrease in sleep latency was discerned within the HFS condition, marking a significant difference from the stim-OFF condition specifically for the light-OFF sessions 2 and 3 (HFS/light-OFF session [1]: 16.6 ± 1.5 min, p = 0.0807; [2]: 11.4 ± 2.1 min, p = 0.0121; [3]: 9.7 ± 1.3 min, p = 0.0106), as depicted in Figure 2b. Both stim-OFF and LFS conditions saw minimal occurrences of sleep episodes, often lasting just a few seconds following sleep onset (stim-OFF/light-OFF session [1]: 0.1 ± 0.1 min; [2]: 0.1 ± 0.1 min; [3]: 0.2 ± 0.1 min). Alternatively, sleep episode durations increased significantly in the HFS condition, showing a marked difference from the stim-OFF condition specifically for light-OFF session 3 (HFS/light-OFF session [1]: 0.6 ± 0.3 min, p = 0.3278; [2]: 1.6 ± 0.5 min, p = 0.1944; [3]: 2.3 ± 0.4 min, p = 0.0444), illustrated in Figure 2c. Over the entire duration of the 3-hour mMSLT procedure, the healthy monkey’s average core body temperature was recorded at 37.7 ± 0.1 °C while it increased to 38.1 ± 0.3 °C under LFS condition yet showed a decrease to 37.3 ± 0.1 °C under the HFS condition, as depicted in Figure 2d. Notably, in both the LFS and HFS conditions, there were observable gradual changes in core body temperature over time (as shown in Figure 2e), with significant differences recorded from the stim-OFF condition during the last 5 minutes of the experiments under LFS condition; however, the HFS condition showed no such difference (illustrated in Figure 2f).
Under the parkinsonian state in the stim-OFF condition, the monkey displayed pronounced sleepiness, indicated by a 100% occurrence rate of at least one sleep episode during each session, faster sleep latency (stim-OFF/light-OFF session [1]: 11.0 ± 2.3 min; [2]: 8.4 ± 2.0 min; [3]: 10.5 ± 2.8 min; mean: 10.0 ± 1.3 min), and extended sleep durations compared to healthy conditions (stim-OFF/light-OFF session [1]: 6.0 ± 1.6 min; [2]: 5.1 ± 1.1 min; [3]: 8.0 ± 1.7 min). Interestingly, the incidence of at least one sleep episode decreased significantly from 100% in the stim-OFF condition to 66% in the LFS condition, while it remained similar in the HFS condition at 83%, as demonstrated in Figure 2g. Within the LFS condition, sleep latency notably increased, achieving statistical significance during light-OFF session 3 when compared to the stim-OFF condition (LFS/light-OFF session [1]: 10.5 ± 1.5 min, p = 0.0998; [2]: 15.5 ± 1.3 min, p = 0.0643; [3]: 18.8 ± 0.8 min, p = 0.0422), as represented in Figure 2h. Furthermore, sleep durations demonstrated a progressive decline within the LFS condition relative to the stim-OFF condition (LFS/light-OFF session [1]: 2.6 ± 0.4 min, p = 0.5232; [2]: 1.5 ± 1.0 min, p = 0.2522; [3]: 0.9 ± 0.9 min, p = 0.0012), illustrated in Figure 2i. Notably, within the HFS condition, there was no significant difference in sleep latency or duration compared to the stim-OFF state. In comparison to the healthy state (37.7 ± 0.1 °C), the parkinsonian state showed a slight reduction in core body temperature (37.4 ± 0.1 °C). Subsequently, core body temperature increased under the LFS condition to 37.7 ± 0.1 °C but tended to see a decrement in the HFS condition down to 37.3 ± 0.1 °C (see Figure 2j). Additionally, core body temperature variations emerged gradually over time in the LFS condition (as represented in Figure 2k), exhibiting significant differences from the stim-OFF condition during the final 5 minutes of experimentation; however, no such variations were recorded within the HFS condition (illustrated in Figure 2l).
Fig. 2
The effects of LHA-DBS at low- and high-frequency stimulation on sleep/wake behavior and core body temperature are summarized as follows: (a, g), Occurrence of at least one sleep episode is expressed as a percentage during all light-OFF sessions combined across stim-OFF (n = 18), LFS-ON (n = 15), and HFS-ON (n = 15) conditions for both healthy (indicated in gray) and Parkinsonian (indicated in orange) states. (b, h), Sleep latency is expressed in minutes ± SEM for each light-OFF session: (1) from 10:00 to 10:20, (2) from 11:00 to 11:20, (3) from 12:00 to 12:20, classified by stim-OFF (n = 6; solid line), LFS-ON (n = 5; bold dotted line), and HFS-ON (n = 5; thin dotted line), for healthy (indicated in gray) and Parkinsonian (indicated in orange) states. c, i, Sleep duration is presented in minutes ± SEM for each light-OFF session: (1) from 10:00 to 10:20, (2) from 11:00 to 11:20, (3) from 12:00 to 12:20, across stim-OFF (n = 6; solid line), LFS-ON (n = 5; bold dotted line), and HFS-ON (n = 5; thin dotted line), with healthy state shown in gray and Parkinsonian state in orange. (d, j), Mean core body temperature is presented in °C ± SEM, compiled during all light-OFF sessions combined in stim-OFF (n = 6), LFS-ON (n = 5), and HFS-ON conditions (n = 5), for healthy (indicated in gray) and Parkinsonian (indicated in orange) states. (e, k), An example illustrates core body temperature trends over time, from 9:30 am to 12:30 pm, under stim-OFF, LFS-ON, and HFS-ON conditions, for healthy (indicated in gray) and Parkinsonian (indicated in orange) states, where black rectangles denote light-OFF sessions. (f, l), Mean core body temperature ± SEM, depicted in °C, recorded during the first and last 5 minutes of experiments in stim-OFF (n = 6), LFS-ON (n = 5), and HFS-ON conditions (n = 5), with healthy state in gray and Parkinsonian state in orange. Statistical significance was determined using the Kruskal–Wallis test followed by Dunn’s multiple comparisons test (p
What are the differences in sleep latency between healthy and parkinsonian states during different stimulation conditions?
Ange) states, with error bars representing standard error of the mean. (b, h), Sleep latency (in minutes) for healthy and parkinsonian states across light-OFF sessions during stim-OFF, LFS, and HFS conditions, highlighting statistically significant differences where applicable. (c, i), Duration of sleep episodes (in minutes) shown for each condition, making comparisons between states clear. (d, j), Average core body temperature (in °C) recorded for healthy and parkinsonian states under different stimulation conditions, with error bars illustrating standard error of the mean. (e, k), Gradual changes in core body temperature over time during stimulation conditions, with significant variations plotted where noted. (f, l), Comparison of temperature measurements during the final 5 minutes of experimentation, illustrating significant differences between conditions where relevant. These insights collectively emphasize the profound impact of low- and high-frequency LHA-DBS on sleep patterns and thermoregulation in both healthy and parkinsonian states, suggesting potential therapeutic avenues for managing sleep disturbances in neurodegenerative conditions.