Optogenetic Frequency Scrambling Of Hippocampal Theta Oscillations Dissociates Working Memory Retrieval From Hippocampal Spatiotemporal Codes Part 2

Łączenie MS optogenetyczne stymulacja i hipokamp wapń obrazowanie

To study the effects of theta manipulations on hippocampal spatialand temporal codes, we combined MS optogenetic stimulation withcalcium imaging in CA1. This experimental paradigm raises twopotentially important issues: GRIN lens implants involve tissue damagethat could alter the physiological state of theta oscillations, and thewavelength spectrum of the excitation LED used to excite GCaMP6fcould potentially overlap with that of the opsin located on the terminalfibers of GABAergic MS fibers in the hippocampus.

hipokamp jest a bardzo ważny struktura w mózgu. Jest głównie odpowiedzialny za przechowywanie i przetwarzanie pamięć informacje. Jako my wszyscy wiemy, pamięć jest ważny część z człowieka intelektualisty działania i ważny sposób dla nas komunikowania i wchodzenia w interakcje z nasze otaczającym środowiskiem. Dlatego, funkcja funkcji hipokampa jest kluczowa do naszego życia.

Hipokamp przestrzeń jest a sposób z opisywany jak przestrzenny informacja jest przetwarzany w nasz mózgach. Odnosi się do aktywnego obszaru a grupy neuronów w nasz mózgu to procesy przestrzenna informacja. Ten obszar jest nazywany przyhipokampowy obszar obszar i jest blisko powiązany z koniem morskim. Badania pokazują to obszar blisko hipokamp procesy informacje powiązane do przestrzeni i pamięć i jest a klucz część nasz pamięć proces.

W szczególności, the hipokamp's przetwarzanie nasz przestrzenny informacja głównie obejmuje dwa aspekty. Pierwszy aspekt jest nasz zmysł kierunku i zdolność do nawigacji. Kiedy my chodzimy, the hipokamp zapisy nasz kroki i pozycja, utrzymywanie nasz orientacja w przestrzeni. Jeśli nasz hipokamp jest uszkodzony, to może powodować problemy takie jako dezorientacja lub bycie niezdolne do znalezienia nasz sposób dom.

Kolejnym aspektem jest pamięć zdolność. Ludzie korzystają the przestrzenna informacja dostarczana przez the hipokamp do pomagają ludzie pamiętaj. Jeśli my doświadczamy coś w a pewne miejsce, the hippocampus będzie przechowywać to informacje, i my możemy dowiedzieć się o tych doświadczeniach lub ludziach poprzez przypominać.

My stale wzmacniamy naszą pamięć możliwości poprzez uczenie się, i hipokamp odgrywa a kluczowa rola w tym procesie. Ponieważ z the importance z the hipokamp, my potrzebujemy z zwracamy uwagę na jego zdrowie i chronić it poprzez niektóre metody. Sudoku, bieganie, uczenie się nowe umiejętności, etc. może wszystko poprawić nasz pamięć i chronić zdrowie nasz hipokamp.

Dlatego, dieta, ćwiczenia, i utrzymanie dobrego snu może pomóc nas chronić nasz hipokamp i poprawiać naszą pamięć zdolność. Kiedy mamy czysty umysł i silna pamięć, możemy lepiej rozumiemy świat i tworzyć a lepsze życie. Może być widzialny to my musimy poprawić naszą pamięć. Cistanche deserticola może znacznie poprawić pamięć, ponieważ Cistanche deserticola może również regulować równowaga neuroprzekaźników, taki taki jak zwiększa poziomy acetylocholiny i czynniki wzrostu . Te substancje są bardzo ważne dla pamięci i uczenia się. W dodatku, mięso może również poprawić krew przepływ i promować tlen dostarczanie, które może zapewnić to dostateczny dostateczny składnikowy i energia, w ten sposób poprawiając mózg witalność i wytrzymałość.

Kliknij wiedz suplementy do do wzmocnienie pamięć

Do zweryfikować czy GRIN soczewka implanty zmienione fizjologia theta, my pierwszy wszczepione my z a GRIN soczewka w prawy hipokampi dwa elektrody w obie lewo i prawo hipokampi, i znalezione porównywalne sygnały theta w w półkulach (ryc. 3a). My porównaliśmy theta oscylacje podczas otwarcia pola eksploracja w my my z oba a GRINlens i an dołączony LFP elektroda, do myszy z elektrody tylko, iznalezione nie znaczące różnice pomiędzy obiema grupami (Rys. 3b). My nie nie znaleźliśmy żadnych znaczących różnic między względnych theta moc wszczepionych myszy z a GRIN soczewka i LFP elektroda (0.14 ± 0.0{07) versus anelektroda tylko (0.154 ± 0.008; t-test, t54=1.060, p=0.29).

Although previous reports have described that combining optogenetic stimulation of ChrimsonR in cell bodies with imaging of GCaMP inneurons at the vicinity of terminals is possible with minimal crosstalk53, wenext monitored any potential opsin activation of MS terminals in the hippocampus by recording CA1-LFP while emitting excitation light with ourminiscope through a GRIN lens. After calibrating mini scope light outputpower (Fig. 3c), we found no effect of the mini-scope blue excitation lighton endogenous theta power (1ANOVA, F(4,295)= 0.7729, p=0.5435; Fig. 3d).Since it has been reported that mini scope excitation light can induce aslight depolarization of terminals transfected with ChrimsonR53, potentiallyhindering further optogenetic-induced depolarization, we next applied MSoptogenetic stimulations while imaging with a ~0.3mW/mm2 mini scope LED power and were able to significantly disrupt or pace theta usingscrambled or 8 Hz stimulation respectively (Friedman test χ2=6.000,p=0.0278; Fig. 3e).

Zakłócenie z theta rytmy modulacje a mała porcja CA1 komórki

We then performed phasic (5 s ON, 5 s OFF) optogenetic stimulationsof MS while recording CA1 pyramidal cells as mice freely explored anopen field (Fig. 4a). We found that a portion of recorded cells wereconsistently excited in these conditions, while others were inhibited(Fig. 4b; see Methods). The activity of pyramidal cells during stimulations was overall lower compared to baseline, for both scrambled stimulation during running (Pearson correlation, R2=0.567, p Less than or equal to 0.00{{20}}1)and rest (R2=0.521, p Less than or equal to 0.0001) periods, as well as for 8 Hz stimulation (R2=0.6, p Less than or equal to 0.0001 for rest periods; R2=0.632, p Less than or equal to 0.0001 for runningperiods; n=1849 cells, N=5 mice; Fig. 4c). Overall, ~6.42 ± 0.52% oftotal cells were significantly modulated by scrambled optogeneticstimulations (Fig. 4d). Among those modulated cells, 50.56 ± 6.38%were inhibited while 49.43 ± 6.38% were excited (n=1849 cells, N=5mice; Fig. 4e).

My następny przeanalizowaliśmy efekty optogenetyczne stymulacja na przestrzenne strojenie z hipokamp neurony jako myszy swobodnie zbadane otwarte pole. to koniec, my my obliczone aktywność szybkość mapy przez używanie epoka w lub zewnętrznie stymulacja okresy (dla linii bazowej warunek, my uwzględnione epoki to następowały takie same 5 s ON, 5 s ON, 5 s OFFpattern used for actual stimulation; Fig. 4f). Stabilność był wtedy obliczony jako korelacja między szybkość mapy dla linia bazowa i stymulacja epoki. Pomimo to wyżej wymienione zmiany w ogólnie aktywność%2skrzynia mapy wyświetlane nie zmiana z przestrzenny stabilność dla albo scrambledor 8 Hz stymulacje (Kruskal–Wallis H3=3.5, p=0.1773; Rys. 4g).

MS optogenetyczne stymulacja zmiany zachowanie ale nie czasoprzestrzenne kody

Do ocena efektów theta zakłócenia na czasowe i przestrzenne kody,we monitorowane CA1 piramidalne neuron aktywność na na the 3-tone liniowe track (Rys. 5a). Tutaj, my dźwignia jeden z główny zalety z obrazowania wapnia, który jest zdolność do rejestracja nagrane komórki ponad kilka dni wykonywanie zakodowane lub 8 Hz stymulacje włączone wybrane dni(Rys. 5b, c, dół panel). Skupiliśmy się nasz analiza na parach z dniz taka sama ilość czasu (48 h) między testowaniem (Rys. 5c, górny panel). dla każdy warunek, my oceniliśmy część z ogółem komórki znacząco kodowanie jedną lub kilka zmiennych i znaleziono nie efekt 8 Hz lub zakodowane stymulacja włączone przestrzenne i czasowe kodowanie(RM-ANOVA; F2=0.807, p=0.453 d główny efekt stymulacji;F6=1.283, p {{15% 7d}.285 dla interakcji pomiędzy stymulacją izakodowaną zmienną, n=5 myszy; Rys. 5d). 

While the portion of cells did not change under stimulation conditions, we assessed the effect of MS stimulation on place-, time-, anddistance-modulated cells' tuning curves stability. To this end, wetracked neurons across days (Fig. 5c; see Methods) and computed thestability of place and time fields as the pairwise correlation betweenfields across 48 h. Because CA1 is known to display prominentremapping over days, we also used a pair of days with no stimulationto compute a baseline stability score to be used as a reference(Fig. 5e–g). We found no change in stability during MS stimulation forplace-modulated cells (1ANOVA, F2=1.907, p=0.1511; n=205 cell pairs pooled from N=5 independent mice; Fig. 5h), time-modulated cells(1ANOVA, F2=2.201, p=0.113; n=227 cell pairs pooled from N=5independent mice; Fig. 5i), and distance-modulated cells (1ANOVA,F2=0.6962, p=0.5024; n=64 cell pairs pooled from N=5 independent mice; Fig. 5j). We also extended the same analysis to conjunctiveneurons (i.e. neurons that can encode more than one variable; Supplementary Fig. 5a–f) and found no effects of optogenetic stimulationson the stability of conjunctive spatial (1ANOVA, F2=3.731, p=0.0661;N=4 independent mice; Supplementary Fig. 5g), temporal (1ANOVA,F2=1.993, p=0.8228; N=4 independent mice; Supplementary Fig. 5h),and distance cells (1ANOVA, F2=0.469, p=0.6400; N=4 independentmice; Supplementary Fig. 5i).

We next assessed the quality of spatiotemporal codes using anaive Bayesian classifier to decode location (Fig. 6a, b), time (Fig. 6cd), and distance traveled (Fig. 6e, f), using random bootstrap samples (n=50 bootstrap samples, n=160 cells per sample). Decodederrors were systematically lower than shuffled surrogates, includingduring 8 Hz or scrambled stimulation for location (2ANOVA,F5=2172, p Less than or equal to 0.0001; n=50 bootstrap samples from one representative mouse; Fig. 6a), time (2ANOVA, F5=1292, p Less than or equal to 0.0001;n=50 bootstrap samples from one representative mouse; Fig. 6c),and distance (2ANOVA, F5=1964, p Less than or equal to 0.0001; n=50 bootstrapsamples from one representative mouse; Fig. 6e) indicating thatspatiotemporal codes were preserved during stimulation. To estimate the inter-individual significance of spatiotemporal codes, wez-scored decoding error by using both the actual and shuffledresults (see Methods) for a given day, for every mouse (Fig. 6b, d, f).MS optogenetic control did not significantly alter the encoding oflocation (1ANOVA, F2,11=2.2332, p=0.1432; N=5 mice; effect sizeη2=0.29; Fig. 6b), time (1ANOVA, F2,11=0.4561, p=0.6452; N=5mice; effect size η2=0.07; Fig. 6d), or distance (1ANOVA,F2,11=0.6102, p=0.5606; N=5 mice; effect size η2=0.09; Fig. 6f).

In the behavioral paradigm used to analyze temporal modulation of neuronal activity, mice are water-scheduled and trained tocollect rewards. Based on this assumption, we quantified the number of returns to an empty reward site as errors, and computed thepercentage of correct trials to total trials as a proxy for performance(Fig. 6g). In these conditions, we found a significant effect ofoptogenetic stimulation on performance across days (Friedman testχ2=6.000, p=0.0278) and in particular a significant difference in performance between during baseline (78.70 ± 2.45%) and scrambled stimulation (44.44 ± 5.56%; multiple comparisons, p=0.0429;N=3 mice; Fig. 6h). Only mice with a minimum of 12 runs wereincluded in this analysis. Because of the limitations of this task (lowcognitive load and a low number of mice tested), we next set out toassess the effect of MS optogenetic stimulation in standardizedmemory tasks.

Zakłócenie theta sygnały upośledzenia przestrzenny rozpoznawanie ipraca pamięć odzyskiwanie

To test the role of theta signals in spatial memory, we used a dedicatedgroup of mice injected with ChrimsonR and implanted with a fiberoptic in the MS (Fig. 7a, left panel). Mice were subjected to a novelplace object recognition (NPOR) task (Fig. 7a, right panel). To assessthe role of theta oscillations in memory encoding and retrieval, weFig. 7|MS optogenetic stimulations disrupt maintenance and retrieval, but notencoding of episodic and working memory. a Mice were implanted with fiberoptics in the MS after transfecting ChrimsonR (top). They were then subjected tothe novel object place recognition task (bottom, see Methods). b In this task, bothscrambled (red) and 8 Hz (blue) stimulations during retrieval, as well as 8 Hz(green) but not scrambled (yellow) stimulations during encoding disrupted memory performance (2ANOVA, F4,31=3.283 for the main effect of treatment,p=0.0097; effect size for the main effect of group, η2p=0.306; N=12 mice). c 

Tofurther examine the effect of MS stimulation on memory encoding, maintenance,and retrieval, mice were trained in a delayed non-match to sample (DNMTS) task inan automated T-maze. d Mice transfected with ChrimsonR (red) or YFP (black) weretrained (in the absence of stimulation) to choose the correct, non-matching armuntil the performance exceeded a criterion of 0.8 portions of correct choicesper day, for at least two consecutive days (green band; RM-ANOVA, F8=8.738,p Less than or equal to 0.0001 for the main effect of training days; pairwise Tukey multiple comparisontests between groups, p=0.420; N=17 mice). e–g Performance during stimulationat different phases of the task for mice injected with ChrimonR (top) or YFP controls (bottom). Red shading indicates the stimulated regions of the maze. e Averagedaily performance when performing MS stimulations during encoding only (RMANOVA, F11=2.197, p=0.547; N=17 mice). f Average daily performance when performing MS stimulations during the 10 s delay period only (RM-ANOVA, F11=3.483,p=0.0495; effect size for the main effect of treatment, η2p=0.109; N=17 mice).g Average daily performance when performing MS stimulations during retrievalonly (RM-ANOVA, F11=3.265, p=0.050; N=17 mice). 

All bar plots and line plotsrepresent the mean ± SEM of at least three independent experiments.Article https://doi.org/10.1038/s41467-023-35825-5Nature Communications|(2023) 14:410 10performed optogenetic stimulations specifically during the sampleand test phases and computed the recognition index (RI; see Methods). When stimulating during retrieval, memory performance wassignificantly reduced for both scrambled (0.472 ± 0.048 RI, n {{10}} mice)and 8 Hz stimulations (0.435 ± {{40}}.057 RI, n=6 mice) groups, comparedto YFP controls that displayed a significant increase in objectexploration during testing (0.61 ± 0.018 RI; 2ANOVA, F4,31=3.283 forthe main effect of treatment, p=0.0097; effect size for the main effectof group, η2p=0.306; N=12 mice; Fig. 7b). On the other hand, scrambled stimulations during encoding did not impair memory at test time(0.60 ± 0.034 RI, p=0.0157, n=6 mice) but 8 Hz stimulation duringencoding lowered memory performance to chance levels(0.39 ± 0.074 RI, p=0.8740, n=6 mice).

To examine the effect of optogenetic control of the MS on specificphases of working memory function (encoding, maintenance, andretrieval), mice were transfected with ChrimsonR and implanted withfiber optics in the MS and trained in a delayed non-match to sample(DNMTS) task. In the sample phase, mice were forced to run to arandomly designated arm to collect a reward. After a delay (10 s), theycould either run in the opposite arm (correct choice) to receiveanother reward or run in the same, unrewarded arm (incorrect arm;Fig. 7c). The advantage of this task is to allow for repetitive testing,specific isolation of task phases (training, delay, testing) and within-subject controls. Mice were trained in this task with no stimulationuntil a criterion performance of 0.8 (a portion of correct trials) wasreached for at least two days. Both ChrimsonR and YFP control micedisplayed significant improvement over time (RM-ANOVA, F8=8.738,p Less than or equal to 0.0001 for the main effect of training days). Importantly, we foundno learning rate differences between the two groups (pairwise Tukey;p=0.420; Fig. 7d). Once mice had learned the rule associated with theDNMTS task, we assessed performance while delivering either scrambled (0.792 ± 0.045) or 8 Hz (0.850 ± 0.036) optogenetic stimulationin the encoding phase (forced choice) only, and did not observe anydifference in performance compared to baseline (0.825 ± 0.030, RMANOVA, F11=2.197, p=0.547, N=17; Fig. 7e). When stimulated onlyduring the delay period, only scrambled stimulations significantlydecreased memory performance (0.733 ± 0.057) compared to baseline(0.825 ± 0.030, RM-ANOVA, F11=3.483, p=0.0495; effect size for themain effect of treatment, η2p=0.109; Fig. 7f). In contrast, when stimulated during retrieval, mice stimulated with 8 Hz displayed significantly reduced memory performance (0.675 ± 0.049) compared tobaseline (0.825 ± 0.030, RM-ANOVA, F11=3.265, p=0.050; Fig. 7g). Incontrast, YFP control mice were not affected by stimulations duringencoding (RM-ANOVA, F2=0.1314, p=0.8781), the delay period (RMANOVA, F2=0.2020, p=0.8197), or retrieval (RM-ANOVA, F2=0.0454,p=0.9557; effect size for the main effect of treatment, η2p=0.196) ofworking memory.

Optogenetyczny kontrola stwardnienie rozsiane neurony nie zmienia porusza się

Importantly, it was previously reported that pacing theta oscillationscould decrease locomotor speed and its variability10, which couldexplain at least in part the effects of optogenetic stimulations onworking and episodic memory. To thoroughly assess the specificity ofMS optogenetic stimulation on memory, we performed additionalexperiments to rule out the direct effects of the optogenetic stimulations on locomotor velocity. A subset of mice injected with ChrimsonRand implanted with fiber optics in the MS as well as LFP electrodes inCA1 were allowed to explore an open field freely while being subjectedto 5 s ON, 5 s OFF optogenetic stimulations (Supplementary Fig. 6a).8 Hz stimulation led to consistent pacing of hippocampal oscillationsto that frequency (Supplementary Fig. 6b). Those stimulations werenot associated with any apparent change in locomotor behavior(Supplementary Fig. 6c), including mean speed (unpaired, two-tailed ttest, t116=0.4140, p=0.6796; Supplementary Fig. 6d) and speedcoefficient of variation (CV; unpaired, two-tailed t-test, t116=0.8296,p=0.4095, n=59 stimulation epochs; Supplementary Fig. 6e). Similarly, scrambled stimulation consistently led to abolished theta oscillations (Supplementary Fig. 6f) but no apparent changes in locomotorbehavior (Supplementary Fig. 6g), including mean speed (unpaired,two-tailed t-test, t118=0.2268, p=0.8210; n=60 stimulation epochs;Supplementary Fig. 6h) and speed CV (unpaired, two-tailed t-test,t118=1.838, p=0.686; n=60 stimulation epochs; Supplementary Fig. 6i).

While natural theta frequency and locomotor speed arecorrelated59, the exact direction of causation between these variablesremains poorly understood. To answer this question, we performedoptogenetic stimulation using a 5s ON, 5s OFF paradigm, and a randomfrequency was selected for each stimulation epoch (SupplementaryFig. 6j). These stimulation frequencies covered the entirety of the thetaband spectrum (Supplementary Fig. 6k). As expected, we found thatnatural theta oscillations frequencies are directly correlated to runningspeed for one example mouse (R2=0.1114, p Less than or equal to 0.0001; SupplementaryFig. 6l). Importantly, when theta oscillation frequency results from MSoptogenetic control, the correlation drops below chance level(R2=0.0006779, p=0.6244; Supplementary Fig. 6m) suggesting thatlocomotion dictates theta frequency, but not the opposite. We systematically replicated these results across mice and observed a drop inthe correlation between theta frequency and locomotor speed underoptogenetic stimulation control (paired t-test, t3=3.922, p=0.0295;N=4 mice; Supplementary Fig. 6n). Altogether, these results supportthat our optogenetic stimulation does not alter locomotor behavior,which is highly relevant for the next behavioral assays.

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