{"artifact":{"id":"experiment_proposal-26b8f40f-b0bc-4eb6-b8dd-5a3bee53aa5e","artifact_type":"experiment_proposal","entity_ids":"[\"h-var-b7e4505525\"]","title":"Experiment Proposal: h-var-b7e4505525","quality_score":0.75,"created_by":"forge_experiment_proposal_generator","provenance_chain":"[]","content_hash":"318ed119940542d2963ad8c5fdc65bcc469bb3194dccfd539181fd6cd2dbd239","metadata":{"citations":["31076275","35151204","36450248","37384704","38642614","39964974","27929004","31578527","39435008","39803521","36513524","28111080","34027028","30155285","36211804","28714589","30936556","33127896","34982715"],"generated_at":"2026-04-28T19:56:43.802436+00:00","intervention":"Closed-loop transcranial focused ultrasound (tFUS) delivered to bilateral entorhinal cortex layer II via a stereotaxically-positioned focused ultrasound transducer (center frequency 650 kHz, ISPTA 300 mW/cm2, 40% duty cycle, 40 Hz burst envelope, 5 ms on/5 ms off); stimulation triggered only when real-time hippocampal LFP gamma power (30-80 Hz) falls below 2 SD of session baseline (sampling rate 1 kHz, Hilbert transform demodulation, 100 ms detection window)","key_controls":["Sham tFUS: identical surgical procedure and EEG implant but no ultrasonic energy delivered (0 mW/cm2)","WT littermate controls: age/sex-matched C57BL/6J x SJL/J F1 mice receiving closed-loop tFUS to EC-II","Off-target stimulation: tFUS delivered to primary somatosensory cortex (S1) at identical parameters to test anatomical specificity","Open-loop tFUS: 40 Hz continuous tFUS without EEG gamma-triggered gating (constant 5-min on/5-min off cycling)","Naive 5xFAD: no surgery, no stimulation, age/sex-matched to control for implant effects","Pre-registered null thresholds: gamma power increase <15% above sham at all timepoints fails primary hypothesis; p-tau ratio change <10% fails secondary pathology criterion"],"model_system":"5xFAD transgenic mice (B6SJL-Tg(APPSwFlLon,PSEN1*M146L*L286V)6799Vas/MuyJ) and age/sex-matched WT littermates; both sexes, 3-4 months of age (early amyloid pathology stage); n=22 per group (estimated 15% attrition yield final n=19)","hypothesis_id":"h-var-b7e4505525","timeline_weeks":28,"_schema_version":1,"null_hypothesis":"Closed-loop transcranial focused ultrasound targeting EC-II SST interneurons produces no measurable restoration of hippocampal gamma oscillation power compared to sham stimulation in the 5xFAD mouse model of Alzheimer's disease","primary_readout":"Hippocampal CA1 stratum radiatum local field potential gamma oscillation power (30-80 Hz) during active navigation (VR running wheel) and slow-wave sleep, expressed as % change from pre-stimulation baseline; spectral analysis via Welch's method (4 s windows, 50% overlap)","estimated_cost_usd":212000.0,"secondary_readouts":["Perforant path input-output function: evoked fEPSP slope (slope 0.1-0.9 ms post-stimulus) in dentate gyrus following angular bundle stimulation at increasing intensities (0-200 uA)","Tau phosphorylation (p-tau S396, S404) and amyloid-beta 42 ELISA from EC and dorsal hippocampus tissue","SST+ interneuron density and c-Fos co-localization in EC layer II via immunohistochemistry","CA1 parvalbumin interneuron firing properties ex vivo (patch-clamp, current-clamp)","Morris water maze spatial navigation accuracy (% time in target quadrant, probe trial)","Theta-gamma coupling strength (phase-amplitude cross-frequency coupling, Modulation Index) in hippocampal LFP","Microglial Prozoning domain 1 (PZ1) marker expression via flow cytometry from EC tissue","P-tau spreading gradient: ratio of EC p-tau to CA1 p-tau (ELISA)"],"expected_effect_size":"Cohen's d = 0.95 for primary gamma power readout; power = 0.80, alpha = 0.05, two-tailed; based on literature-reported effect of 40 Hz gamma entrainment on hippocampal LFP power in AD models (PMID 38642614) and estimated 40-50% restoration relative to WT baseline","statistical_approach":"Linear mixed-effects model (lme4 package, R) for repeated-measures primary readout with fixed effects: treatment, time, genotype, sex; random intercepts for animal ID; planned contrasts: closed-loop tFUS vs sham (primary), closed-loop vs open-loop (mechanistic), 5xFAD-WT comparison (validation); alpha = 0.05; Bonferroni correction for 7 secondary readouts (corrected alpha = 0.007); effect sizes reported as marginal R-squared; Shapiro-Wilk normality test and Levene's variance homogeneity test applied prior to model fitting; pre-registered analysis plan deposited on Open Science Framework","sample_size_rationale":"Power analysis (G*Power 3.1): two-group comparison (tFUS vs sham), two-tailed t-test, effect size d=0.95, alpha=0.05, power=0.80 yields n=18/group; adding 15% attrition buffer gives n=21/group; inflated to n=22 for balanced sex distribution; total N=110 (5 groups x 22); rationale based on gamma entrainment literature in AD mice (PMID 38642614) and tFUS dose-response studies in related neuromodulation protocols","falsification_criterion":"Primary: Closed-loop tFUS group fails to demonstrate statistically significant increase in hippocampal gamma power above sham-stimulation baseline at any post-stimulation timepoint (weeks 2, 4, 6); effect size 95% CI includes zero; Secondary: No rescue of spatial memory (MWM target quadrant time <25% during probe trial); Secondary: No restoration of perforant path input-output gain (max slope within 1 SD of 5xFAD sham group); Secondary: p-tau EC/CA1 ratio unchanged from sham; Tertiary: Any tFUS group shows >2x increase in c-Fos+ neuron density outside target region or spontaneous seizure occurrence (EEG spike frequency >3 SD above baseline) indicating off-target hyperexcitability; Falsification at interim analysis (week 12, n=8/group) triggers immediate study halt"},"created_at":"2026-04-28T12:56:43.818935-07:00","updated_at":"2026-04-28T12:56:43.818935-07:00","version_number":3,"parent_version_id":null,"version_tag":null,"changelog":null,"is_latest":1,"lifecycle_state":"active","superseded_by":null,"deprecated_at":null,"deprecated_reason":null,"dependencies":null,"market_price":0.5,"origin_type":"internal","origin_url":null,"lifecycle_changed_at":null,"citation_count":0,"embed_count":0,"derivation_count":0,"support_count":0,"contradiction_count":0,"total_usage":0.0,"usage_score":0.5,"usage_computed_at":null,"quality_status":null,"contributors":[],"answers_question_ids":null,"deprecated_reason_detail":null,"deprecated_reason_code":null,"commit_sha":null,"commit_submodule":null,"last_mutated_at":"2026-05-16T14:51:34.657673-07:00","disputed_at":null,"gap_id":null,"mission_id":null,"intrinsic_priority":null,"effective_priority":null,"artifact_id":"e8d9247f-a37d-4c75-a60e-5b69bca4753c","artifact_dir":null,"primary_filename":null,"accessory_filenames":null,"folder_layout_version":1,"migrated_to_folder_at":null,"hypothesis_id":null,"authorship":{"kind":"human","contributors":[{"role":"author","actor_ref":"forge_experiment_proposal_generator"}]},"epistemic_tier":"T3_provisional","created_by_agent_id":null},"outgoing_links":[{"target_artifact_id":"h-var-b7e4505525","link_type":"derives_from","strength":1.0,"evidence":"{\"source\": \"forge_experiment_proposal_generator\", \"method\": \"generate_experiment_proposal\"}"}],"incoming_links":[],"current_artifact_id":"experiment_proposal-26b8f40f-b0bc-4eb6-b8dd-5a3bee53aa5e","is_canonical":true,"supersede_chain":["experiment_proposal-26b8f40f-b0bc-4eb6-b8dd-5a3bee53aa5e"]}