Persistent Brain Connectivity Changes in Healthy Volunteers Following Nitrous Oxide Inhalation

Abstract

Background

Nitrous oxide holds promise in the treatment of major depressive disorder. Its psychotropic effects and NMDA receptor antagonism have led to comparisons with ketamine. Despite longstanding use, persistent effects of nitrous oxide on the brain have not been characterized.

Methods

Sixteen healthy volunteers were recruited in a double-blind crossover study. In randomized order, individuals underwent a 1-hour inhalation of either 50% nitrous oxide/oxygen or air/oxygen mixtures. At least two 7.5-minute echo-planar resting-state functional magnetic resonance imaging scans were obtained before and at 2 and 24 hours after each inhalation (average 130 min/participant). Using the time series of preprocessed, motion artifact–scrubbed, and nuisance covariate–regressed imaging data, interregional signal correlations were measured and converted to T scores. Hierarchical clustering and linear mixed-effects models were employed.

Results

Nitrous oxide inhalation produced changes in global brain connectivity that persisted in the occipital cortex at 2 and 24 hours postinhalation (p < .05, false discovery rate–corrected). Analysis of resting-state networks demonstrated robust strengthening of connectivity between regions of the visual network and those of the dorsal attention network, across 2 and 24 hours after inhalation (p < .05, false discovery rate–corrected). Weaker changes in connectivity were found between the visual cortex and regions of the frontoparietal and default mode networks. Parallel analyses following air/oxygen inhalation yielded no significant changes in functional connectivity.

Conclusions

Nitrous oxide inhalation in healthy volunteers revealed persistent increases in global connectivity between regions of primary visual cortex and dorsal attention network. These findings suggest that nitrous oxide inhalation induces neurophysiological cortical changes that persist for at least 24 hours.

Nitrous oxide is an odorless, colorless gas that has been routinely employed in clinical practice for over 150 years due to its analgesic, amnestic, and hypnotic properties. Its psychomimetic effects and noncompetitive antagonism of NMDA receptors (NMDARs) have led to comparisons with ketamine (1,2). Recently, nitrous oxide has shown promise in the treatment of major depressive disorder (3, 4, 5, 6), posttraumatic stress disorder (7), alcohol withdrawal (8), and bipolar depression (9,10). Despite widespread and longstanding use, mechanistic understanding of nitrous oxide’s effects on the brain have not been fully elucidated.

Persistent effects on brain activity (e.g., improved mood) occurring with brief exposure to nitrous oxide are thought to mirror those of ketamine, i.e., they are rapid in onset and sustained for a week or more. Two studies have now demonstrated that antidepressant effects of a single nitrous oxide inhalation can persist for at least 1 to 2 weeks (3,4). These persistent effects parallel those observed with ketamine; a sustained antidepressant benefit for days after a single ketamine exposure has been a reproducible finding, with peak effects at 24 hours after administration (11). The persistent antidepressant effects of ketamine are attributed in part to synaptic plasticity within networks of brain regions. For ketamine, functional neuroimaging has revealed sustained changes in brain connectivity in individuals with depression (12, 13, 14, 15, 16, 17, 18), including visual (18,19) and motor cortical areas (17). Similarly, healthy volunteers also demonstrated changes in cortical connectivity in a double-blind crossover study (20). It is not known whether nitrous oxide induces similar plasticity changes in brain connectivity in individuals with or without depression.

Neuroimaging studies of the acute effects of nitrous oxide inhalation have shown altered neural activity across distributed brain regions (21, 22, 23). Inhaled nitrous oxide increases cerebral blood flow in occipital regions (24) and activation in the anterior cingulate cortex but bilateral deactivation in the posterior cingulate, hippocampus, parahippocampal gyrus, and visual association areas (25). In contrast, there have been no nitrous oxide neuroimaging studies conducted to determine whether such changes persist after inhalation. Such investigations may help identify the loci of action of nitrous oxide’s persistent antidepressant effects, which extend up to weeks after a single nitrous oxide inhalation (3,4).

Here, we conducted a double-blind crossover investigation of subacute (2 hours) and persistent (24 hours) effects of nitrous oxide on cortical functional connectivity, using resting-state functional magnetic resonance imaging (rs-fMRI) of healthy volunteers without depression.

Methods and Materials

Participants and Study Design

The parent study was registered (ClinicalTrials.gov; Identifier NCT02994433) and approved by the Washington University School of Medicine Human Subjects Human Research Protection Office. Data from the depressed cohort will not be presented here.

Following eligibility screening, 21 healthy participants were recruited throughout the greater St. Louis, Missouri, community and through the Volunteer for Health registry at Washington University School of Medicine in St. Louis. All participants fulfilled the following inclusion criteria: age 18 to 65 years, right-handed, good command of English, no history of depression as determined by structured clinical interview [the Mini-International Neuropsychiatric Interview (26)], and not currently depressed [i.e., scored 7 or lower on the 17-item Hamilton Depression Rating Scale (27)]. Additionally, participants did not meet any of the following exclusion criteria: a primary DSM-IV Axis I diagnosis as documented in medical records and as determined by structured clinical interview; known primary neurological disorders or medical disorders impacting brain imaging including dementia, stroke, encephalopathy, Parkinson’s disease, brain tumors, multiple sclerosis, seizure disorder, or severe cardiopulmonary disease; central nervous system active medications as determined by study investigator; known disease affecting drug metabolism and excretion (e.g., renal or liver disease), as determined by study investigator; ineligibility for MRI scans (e.g., a history of claustrophobia or MRI-incompatible metal implanted); current use of psychotropic medications, antidepressants, or prescription or nonprescription drugs/herbals intended to treat depression or anxiety; recent (within the past 12 months) history of substance dependence or abuse, as determined by reported history or urine drug screen; or ability to become pregnant and not using effective contraception. In addition, participants could not have contraindications against nitrous oxide exposure or study procedures: pneumothorax, bowel obstruction, middle ear occlusion, elevated intracranial pressure, chronic cobalamin and/or folate deficiency treated with folic acid or vitamin B₁₂, pregnancy or breastfeeding, inability to provide informed consent, or any other factor that in the investigators’ judgment might have affected patient safety or compliance (e.g., distance greater than 100 miles from the clinic). Written informed consent was obtained from all study participants prior to undergoing any trial activities. Participants received remuneration for their time and effort.

The study design was a prospective, placebo-controlled, double-blind crossover study with 5 study visits (Figure 1). Inhalation order was randomized for the 2 arms (intervention vs. placebo) using a random number generator (block size of 2). The sessions were 4 to 6 weeks apart to minimize carryover effects between conditions that have been observed previously in nitrous oxide inhalation studies (3). Six individuals withdrew after the first inhalation session or the first postinhalation imaging session (CONSORT [Consolidated Standards of Reporting Trials] flow diagram in Figure S1). Recruitment from October 2015 to February 2020 was halted during the COVID-19 pandemic; imaging and inhalation facilities were closed to investigation for an extended period. Despite completing all inhalation and imaging sessions, 1 individual was excluded due to use of a different scanner system and imaging protocol. The final sample (N = 14) included 7 women, with a median age of 44.5 years. When self-identified by race and ethnicity, 2 (14%) were Asian, 5 (36%) were Black, 7 (50%) were White, and 1 (7%) was Hispanic. No participants had active or recent tobacco use. Participant racial and ethnic demographics are included in Table S1. Table S2 lists concurrent medications, medical comorbidities, and adverse events encountered during inhalation sessions.

Figure 1.

Study design. Study participants were randomized to a double-blind crossover study with 60-minute inhalation sessions of 50% nitrous oxide (N₂O)/oxygen and air/oxygen mixtures. Inhalations were separated by a washout period of 4–6 weeks. Resting-state functional magnetic resonance imaging scanning was performed 1–2 hours preinhalation, 2 hours postinhalation, and approximately 24 hours postinhalation. Schematic created using BioRender.

For the intervention condition, participants inhaled 50% nitrous oxide (balance air/oxygen) for 1 hour. We utilized a dose of nitrous oxide that had conferred an antidepressant response in a previous trial, 50% inhalation for 1 hour (3). In the placebo condition, subjects inhaled air/oxygen mixture for 1 hour. Except for the composition of the gas mixtures, inhalations were identical. A face mask and strap were used to generate a semiclosed system connected to an anesthesia machine or a U.S. Food and Drug Administration–approved Porter/Praxair MXR breathing circuit. Total gas flow was kept at 2 to 8 L/min, with nitrous oxide concentrations increased gradually over the first 10 minutes of the inhalation. Participants were monitored according to the American Society of Anesthesiologists standards, including an available attending anesthesiologist, 3-lead electrocardiography, pulse oximetry, end-tidal carbon dioxide, and noninvasive measurement of blood pressure. Following inhalation, subjects underwent a 1-hour observation period for monitoring vital signs and potential adverse events.

Extent of blinding was dependent on the role in the study. The anesthesia team conducted the inhalation per treatment arm assignment. Concealment was undertaken by shielding the study gas flowmeters from the study participant and by recusal of the assessor during the inhalation. Thus, blinding to the inhalation conditions was maintained for participants, the psychiatry team performing the ratings, and the fMRI data analyst. To evaluate the effectiveness of the blinding, participants were assessed with a 5-point Likert scale in which they were asked to rate the extent to which they knew that they had inhaled nitrous oxide (“strongly believe nitrous oxide,” “somewhat believe nitrous oxide,” “somewhat believe placebo,” “strongly believe placebo,” and “don’t know”).

rs-fMRI Data Acquisition

Images were acquired using a Siemens 3T Prisma scanner with a 64-channel head coil. At each imaging time point (Figure 1), at least two 7.5-minute rs-fMRI scans were acquired preinhalation (baseline), 2 hours postinhalation, and approximately 24 hours postinhalation (range 22–28 hours afterward), for a total of 6 scanning sessions. We acquired imaging at 2 and 24 hours following inhalation because antidepressant responses were observed at these times in a previous trial (3). Blood oxygen level–dependent (BOLD) echo-planar imaging was acquired (2.4 mm³ voxels, echo time = 30 ms, repetition time = 0.8 seconds, 552 volumes/scan). During all resting-state scans, participants were instructed to keep their eyes open while focusing on a fixation cross. They were told that they could relax but should remain awake. Compliance with this instruction was not monitored. FIRMM (Frame-wise Integrated Real-time MRI Monitoring; https://turingmedical.com/) software was used to assist in evaluating head motion during data acquisition. Four foam pads were utilized to minimize head movement during imaging. Whole-head T1-weighted structural images (1 mm³ voxels) were also acquired during the preinhalation session.

Image Analysis Preprocessing

Preprocessing and subsequent analyses were performed using the Conn toolbox (28). rs-fMRI interregional correlations were calculated from BOLD image time series that had been slice time corrected, realigned, coregistered with the high resolution T1-weighted volume, spatially normalized, and spatially smoothed with an 8-mm kernel. The beginning and end of the time series was temporally smoothed with a digital Hanning filter to reduce potential edge effects (28,29). This preprocessing incorporated a quality assurance protocol that excluded outlier time points, i.e., those that were >3 standard deviations from the mean global signal intensity or exhibited >0.3-mm scan-to-scan head motion. Imaging quality statistics are included in Table S3. Participants had a median imaging time of 131 minutes (9827 imaging volumes) across the 6 imaging sessions. Physiological sources of noise, estimated from gray and white matter segments using anatomical CompCor (30), were treated as confounds. Time series characterizing estimated subject motion, 6 parameters representing their first-order temporal derivatives, scan-to-scan motion outliers, and the BOLD-contrast signal within the subject-specific white matter and cerebrospinal fluid masks were used as nuisance variables. The resulting residual time series were bandpass filtered from 0.008 to 0.20 Hz. Interregional correlations were then computed and converted to T scores.

First-Level Modeling

Voxel-to-voxel (local correlation [LCOR] and global correlation [GCOR]) and interregional (32 regions comprising the canonical networks) (Table S4) correlations were computed to identify areas showing changes in functional connectivity after nitrous oxide inhalation.

Local connectivity strength was estimated using the LCOR metric (31). LCOR was computed as the average of correlation coefficients between each individual voxel and its neighboring voxels. LCOR maps demonstrate the estimated average correlation strength and sign between a voxel and the neighboring areas in the brain.

Global connectivity strength was estimated using the GCOR metric. For a single voxel in space, GCOR is the average of correlation coefficients between that voxel and all voxels across the brain (32). GCOR provides a measure of node centrality operationalized as the strength and sign of connectivity between a given voxel and the rest of the brain.

Region-based functional connectivity was subsequently used to characterize the spatial pattern of connectivity changes associated with resting-state networks. These estimates involved computing interregional correlations among the regions (Table S4) that were then converted to T scores. A hierarchical clustering approach was then used to identify clusters with overall changes across conditions. To mitigate bias introduced by serial testing, false discovery rate corrections for multiple tests were employed (33).

Second-Level Modeling

We estimated the effects of nitrous oxide on neural activity at a group level, leveraging maps of GCOR and LCOR obtained from first-level modeling. A repeated measures mixed-effects model was implemented, with fixed effects of replication, session, condition, and subject as a random effect. The effect of replication accounted for any differences in effects among the 3 scans that comprised each session of the study arm; the model included coding of session (−1, 0, +1). The effect of session examined linear effects among the imaging days that comprised the experiment. The linear effect of condition allowed the contrast of preinhalation to postinhalation data in the different sessions.

The principal contrasts compared the pre-nitrous oxide condition to the post-nitrous oxide condition at both 2 and 24 hours postinhalation. Parallel analyses utilized this approach for the pre-air/oxygen inhalation and post-air/oxygen inhalation imaging data that were acquired at similar time points.

Results

Study Outcomes

We assessed the effectiveness of masking our treatment conditions across the 28 inhalation sessions. Subjects were asked whether they had received nitrous oxide or placebo. Participants correctly guessed the condition in 21 of 28 sessions (75%) but assumed incorrectly in 5 of 28 sessions (18%) or were unable to ascertain after 2 of the inhalations (7%).

Global Connectivity Following Nitrous Oxide Inhalation in Visual Cortical Regions

Using voxel-based GCOR analyses, voxels clustered within the occipital cortex demonstrated stronger correlations across the brain following nitrous oxide inhalation (Figure 2A, B). No voxels displayed a reduction in correlation (data not shown). These changes in the calcarine sulcus were present at 2 hours and persisted at 24 hours postinhalation (p < .05 for the linear relationship across time) (Figure 2C). A parallel pre-post analysis of the air/oxygen mixture inhalations revealed no effects at the same multiple-test–corrected critical threshold (data not shown). Thus, across the brain, visual cortex voxels showed the greatest change in correlated brain activity with persistence at 24 hours following nitrous oxide inhalation but not placebo.

Figure 2.

Brain-wide global correlation (GCOR) maps identify bilateral primary visual cortex as regions with subacute (2 hours) and persistent (24 hours) changes in GCOR strength following nitrous oxide (N₂O) inhalation. (A) Axial/transverse sections demonstrating occipital gray matter regions with significant increases in GCOR (pre- vs. postinhalation of N₂O). (B) Posterior perspective of surface map for significant increases in GCOR 24 hours post-N₂O inhalation. (C) Bar graph demonstrating cluster mean effect sizes for GCOR at preinhalation and postinhalation (at 2 and 24 hours). Effect sizes are shown with 95% CIs. A positive linear relationship between GCOR and time was observed (cluster k = 228 voxels; p < .05 with false discovery rate multiple-test correction).

Local Connectivity Following Nitrous Oxide Inhalation

Using voxel-based LCOR analyses, no suprathreshold voxels were detected following nitrous oxide inhalation (voxel threshold p < .001 [uncorrected]; cluster threshold p < .05 familywise error rate–corrected). A parallel analysis of the air/oxygen mixture revealed no effects at the same multiple-test–corrected critical threshold (data not shown).

Functional Connectivity Among Canonical Networks Following Nitrous Oxide Inhalation

Additional analyses were undertaken to characterize changes in connectivity across canonical resting-state networks represented by 32 cortical regions (Table S4). As a quality control, preinhalation imaging verified established connectivity patterns across resting-state networks (Figure S2). Using these same 32 cortical regions, pre-post nitrous oxide inhalation connectivity analysis demonstrated that regions of the visual network and dorsal attention network showed stronger correlated activity at both 2 and 24 hours (Figure 3), which withstood false discovery rate correction for multiple comparisons during network-based statistical analysis. Linear increases in functional connectivity were noted among visual cortical regions, as well as between the visual network and the dorsal attention network regions in the intraparietal sulcus and frontal eye fields. Less pronounced connectivity changes were noted between the visual cortical regions and lateral prefrontal cortical regions of the frontoparietal network, the posterior cingulate cortex, and the left lateral parietal region of the default mode network. Small connectivity changes were also observed between the visual cortex and regions of the sensorimotor network, regions in the language network, and posterior regions of the cerebellum. Lastly, weaker correlations were noted between the medial visual cortical region and the posterior parietal cortex of the frontoparietal network. A correlation matrix of interregional changes across all 32 cortical regions is provided in Figure S3. A parallel analysis of the air/oxygen mixture (placebo condition) data revealed no effects at the same multiple-test–corrected critical threshold (data not shown). Thus, following nitrous oxide, but not placebo inhalations, persistent changes in network connectivity were identified linking primary and higher-order visual processing regions and visual attention areas and secondarily to motor and language networks.

Figure 3.

Connectome ring demonstrating functional connectivity changes at 2 and 24 hours after inhalation of nitrous oxide (N₂O). (A) Connectome ring indicating strength of correlations among regions of respective resting-state networks showing linear changes in connectivity across the 3 conditions of preinhalation, 2 hours postinhalation, and 24 hours postinhalation of N₂O (threshold-free cluster enhancement = 55.94, p < .05). Red/yellow color show stronger correlations whereas blue colors indicate weaker correlations within the cluster. Inset shows locations of visual regions. (B) Effect sizes and 95% CIs for changes in connectivity across conditions for the cluster shown in (A). Resting-state networks: Sensorimotor, somatosensory motor network; Visual, visual network; DorsalAttention, dorsal attention network; Cerebellar, cerebellum; DefaultMode, default mode network; Frontoparietal, frontoparietal network; Language, language/auditory network; Salience, salience network. Individual regions are listed in Table S4. ACC, anterior cingulate cortex; FEF, frontal eye fields; IFG, inferior frontal gyrus; IPS, intraparietal sulcus; l, left; LP, lateral parietal; LPFC, lateral prefrontal cortex; MPFC, medial prefrontal cortex; PCC, posterior cingulate cortex; PPC, posterior parietal cortex; pSTG, posterior superior temporal gyrus; r, right; RPFC, lateral rostral prefrontal cortex; SMG, supramarginal gyrus.

Discussion

Here, we demonstrated, in a group of normal control participants without depression, sustained brain network changes following nitrous oxide inhalation. Using a data-driven approach, we observed increased global connectivity within the bilateral calcarine sulci of the occipital lobe. In an independent analysis, a similar cluster of visual regions demonstrated a linear increase in connectivity from 2 to 24 hours postinhalation, with the strongest increases in connectivity to dorsal attention network regions. Weaker changes in connectivity were noted between the visual cortex and regions of the frontoparietal and default mode networks. In contrast, parallel analyses for imaging data obtained from the same individuals after inhalation of air/oxygen mixture did not recapitulate these findings. To our knowledge, this is the first evidence demonstrating sustained alterations in network connectivity following nitrous oxide inhalation, i.e., the effects persisted for at least 24 hours after inhalation of nitrous oxide.

Our findings are consistent with the limited number of previous reports of persistence of either brain imaging or behavioral changes following nitrous oxide inhalation. A positron emission tomography crossover study of 10 individuals evaluated changes in global and regional cerebral metabolic rateduring and after inhalation of 50% nitrous oxide or 30% oxygen (34). While no changes in global cerebral metabolic rate (CMR) were observed during nitrous oxide inhalation, regional CMR was augmented by 14% in the basal ganglia and 22% in the thalamus. Persistent augmentation in cortical CMR was observed in a subset of participants who were imaged 1 hour after crossing over from a previous inhalation of 50% nitrous oxide. More specifically, sustained effects in frontal, temporal, parietal, and occipital cortical regions contrasted with an absence of change in the cerebellum, thalamus, or basal ganglia. Analogous observations of nitrous oxide effects on the electroencephalogram (EEG) have been reported, with increases in frontal theta (35) or delta (36) EEG power above baseline 15 minutes following cessation of nitrous oxide inhalation. Because these studies only measured for 15 minutes after nitrous oxide inhalation, it is unknown whether these EEG changes persist for 2 hours or for 24 hours, consistent with our imaging findings. From a behavioral standpoint, the persistence of augmented cortical CMR is consistent with a small (N = 12) randomized double-blind healthy volunteer study comparing sleepiness following 30-minute inhalations of 40% nitrous oxide, 20% nitrous oxide, or 100% oxygen (37). After inhalation of 40% nitrous oxide, participants were less sleepy at 1 hour as assessed by the multiple sleep latency test compared with those who inhaled 0% or 20% nitrous oxide. In contrast, no changes in performance were observed on 5 psychomotor tests following any of the 3 inhalation conditions. A rapid recovery in cognitive task performance following inhalation was also observed in another study evaluating recall, reaction time, and attention (38). These data are consistent with resolution of subjective mood, memory impairment, and psychomotor impairment in healthy volunteers within 20 minutes of cessation of nitrous oxide (39). However, one small open-label study (N = 7) reported persistent impairment of free recall at 20 minutes following nitrous oxide and an augmented score of pleasantness at 24 hours (40). Thus, our data advance spatial and temporal characterization of neural changes that persist beyond the pharmacokinetic elimination of nitrous oxide.

Given the similarities in purported mechanisms of action for nitrous oxide and ketamine (for example NMDAR antagonism), these nitrous oxide data warrant comparison with sustained brain effects observed following ketamine exposure. Several findings in this study are consistent with previous data demonstrating enhanced synaptic plasticity in visual cortical areas following administration of ketamine. Sumner et al. (41,42) analyzed visual evoked EEG responses in patients with treatment-resistant depression 3 to 4 hours after intravenous subanesthestic ketamine or placebo (remifentanil) (41), evaluating changes in evoked potentials as a marker of synaptic change. Components of the visual evoked response were enhanced by periodic visual stimulation, with persistent enhancement only after ketamine. This study suggested significant modulation of EEG potentials at the bilateral inferior temporal cortex and superior parietal cortex evoked by bilateral middle occipital gyrus sources over a time course that is consistent with our current neuroimaging results. These changes in processing deviant/novel stimuli may also extend to auditory stimuli (42). Parallels between nitrous oxide and ketamine are limited, however, because NMDAR antagonism is only one molecular mechanism of action for these two agents that have many other targets (e.g., GABA [gamma-aminobutyric acid], nicotinic acetylcholine, and muscarinic acetylcholine receptors) (43). Furthermore, actions beyond NMDAR may be critical to the antidepressant effects of both anesthetic agents (44,45).

We cautiously speculate on how brain connectivity alterations following nitrous oxide inhalation in individuals without depression may identify potential loci of action that would underlie antidepressant response. Aberrant functional connectivity within and between resting-state networks has been associated with depression (46), with hyperconnectivity within and between the frontoparietal and default mode networks and hypoconnectivity between the frontoparietal and dorsal attention networks. Nitrous oxide–mediated enhancement of connectivity to nodes with aberrantly strong interconnectivity could conceivably disrupt pathologic hyperconnectivity. More broadly, a diffuse enhancement of synaptic plasticity could contribute to a reversal of pathologic hypoconnectivity. The lack of any modulation of connectivity to the salience network would seem to suggest that action at this network is less likely to be observed in imaging investigations of individuals with depression who are treated with nitrous oxide. These lines of conjecture warrant evaluation in future studies to also address whether these persistent changes in brain connectivity emerge after 25% inhalation of nitrous oxide in the same manner as antidepressant effects (4).

It is important to consider the limitations and strengths of this work. The current study is limited by a small sample size, which is a continued area of methodological consideration (47). This precludes subgroup analyses on sex or other factors. However, this study did include up to 30 minutes of BOLD resting-state scanning time for each scan session, which is considerably longer than in most trials. In addition, the lack of an active placebo must be acknowledged (48). Furthermore, extrapolating our findings from healthy control participants without depression to patients with depression is not unprecedented (20) but should be done cautiously because drug effects on the two populations may be disparate (49). Because we did not image individuals during inhalation, we are not able to ascertain whether functional connectivity changes emerged during inhalation or postinhalation (rebound). Finally, this study was focused on resting-state wakefulness, i.e., there is a lack of behavioral tasks to correlate with these imaging findings. These weaknesses are offset by the crossover design of the study, with individuals serving as their own controls.

These hypothesis-generating data predict that there may be an emergence of functional changes in visual attention and processing in healthy participants. It is not yet clear whether these connectivity changes are related to antidepressant effects or perhaps represent another behavioral change, e.g., increased alertness/reduced sleepiness as previously described (34). Studies are underway to determine whether the observed changes in brain connectivity also manifest in individuals with clinical depression/treatment-resistant depression and whether there are associated changes in depressive symptom severity. Although preliminary, these findings suggest that a rapid and persistent effect on intracortical connectivity could potentially contribute to antidepressant effects; i.e., the patient with depression, by undergoing alteration in brain activity in the visual and attention regions, may “see” and interpret the world differently. In parallel, these data may point to alternative ways to consider depression circuitry that may be relevant to psychiatric therapeutics.