Distinctive Molecular and Metabolic Profiles of 
Chemically Synthesized Psilocybin and Psychedelic Mushroom Extract

doi:10.21203/rs.3.rs-3146433/v1

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Distinctive Molecular and Metabolic Profiles of Chemically Synthesized Psilocybin and Psychedelic Mushroom Extract

https://doi.org/10.21203/rs.3.rs-3146433/v1

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published 20 Feb, 2024

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Psilocybin, a naturally occurring, tryptamine alkaloid prodrug, is currently being investigated for the treatment of a range of psychiatric disorders. Preclinical reports suggest that the biological effects of psilocybin-containing mushroom extract or “full spectrum” (psychedelic) mushroom extract (PME), may differ from those of chemically synthesized psilocybin (PSIL). We compared the effects of PME to those of PSIL on the head twitch response (HTR), neuroplasticity-related synaptic proteins and frontal cortex metabolomic profiles in male C57Bl/6j mice. HTR measurement showed similar effects of PSIL and PME over 20 minutes. Brain specimens (frontal cortex, hippocampus, amygdala, striatum) were assayed for the synaptic proteins, GAP43, PSD95, synaptophysin and SV2A, using western blots. These proteins are indicators of synaptic plasticity. Three days after treatment, there was minimal increase in synaptic proteins. After 11 days, nested analysis of variance (ANOVA) showed a significant increase in each of the 4 proteins over all brain areas studied for PME versus vehicle control, while significant PSIL effects were observed only in the hippocampus and amygdala and were limited to PSD95 and SV2A. Metabolomic analyses of the pre-frontal cortex were performed by untargeted polar metabolomics utilizing capillary electrophoresis – Fourier transform mass spectrometry (CE-FTMS) and showed a differential metabolic separation between PME and vehicle groups. The purines guanosine, hypoxanthine and inosine, associated with oxidative stress and energy production pathways, showed a progressive decline from VEH to PSIL to PME. In conclusion, our synaptic protein findings suggest that PME has a more potent and prolonged effect on synaptic plasticity than PSIL. Our metabolomics data support a gradient of effects from inert vehicle via chemical psilocybin to PME further supporting differential effects. Further studies are needed to confirm and extend these findings and to identify the molecules that may be responsible for the enhanced effects of PME as compared to psilocybin alone.

Biological sciences/Neuroscience

Biological sciences/Molecular biology

psychedelics

psychedelic mushrooms

psilocybin

synaptic proteins

metabolomics

There is renewed interest in studying psychedelics for a number of psychiatric disorders including depression, PTSD, obsessive-compulsive disorders and anxiety ¹. To date, clinical trials have generally been conducted with chemically synthesized psilocybin (PSIL), and little attention has been given to additional, potentially therapeutic, psychoactive or non-psychoactive compounds found in psychedelic mushrooms ². The same is true for other psychedelics that are currently a focus of clinical research. Dimethyltryptamine (DMT) is of plant and animal origin and has also been identified in humans ³. 5-methoxy-dimethyltryptamine (5-MeO-DMT) is derived from the incilius alvarius species of toad ⁴ and the most common source of mescaline is the peyote cactus ⁵. LSD was originally synthesized from lysergic acid which is derived from ergotamine, an alkaloid found in the wheat-infecting ergot fungus. ⁶. In most clinical studies chemically synthesized forms of these compounds are used. Naturally occurring psychedelic compounds do not exist in isolation but are produced by the organism as part of an extensive milieu that includes molecules of many different types. It is possible that these molecules exert significant biological effects or modulate the action of the active molecule in different ways. In regard to cannabis, it is recognized that not only cannabidiol, but a number of other “entourage” molecules exert significant biological effects along with those of delta-9-tetrahydrocanabinol (Δ -9 THC) and modify the effect of Δ − 9 THC ⁷.

Many experienced psychedelic mushroom users believe that different Psilocybe species have distinctive and characteristic effects ^{8, 9}. These reports are consistent with observations regarding the effects of, Inocybe aeruginascens (high in aeruginascin) as compared to the effects of mushrooms with a high psilocybin and psilocin content. Gartz ¹⁰ observed an increased mood-enhancing effect of mushrooms high in aeruginascin as compared to those high in psilocybin, leading him to propose an "entourage effect" of psychedelic mushrooms, whereby additional components of the mushroom extract enhance the effect of psilocybin. A preclinical study compared the effects of an extract from Psilocybe argentipes to chemical psilocybin on marble-burying behavior in mice (an animal screening model used to study OCD) ¹¹. The results of the study showed that the psilocybin mushroom extract was more effective in reducing marble-burying than chemical psilocybin, doing so at a dose of 0.25 mg/kg psilocybin as compared to chemical psilocybin which required 1.5 mg/kg and thus supporting further exploration of an “entourage effect”. Further evidence for a psychedelic mushroom “entourage effect” was provided by Zhuk et al. ¹², who showed that Psilocybe semilanceata and Pholiotina cyanopus mushroom extracts were significantly more potent on in-vivo serotonin 5-HT2A receptor behavioral assays (as demonstrated by the head-twitch response in mice) than the same dose of pure psilocin (the active metabolite of psilocybin).

The variability in effects produced mushroom extracts compared to chemical psilocybin may be explained by differing psilocybin content ¹³ as well as by the presence and varying levels of other potentially bioactive compounds in the different species of mushrooms. These include tryptamines, such as 4-phosphoryloxy-N-methyltryptamine (baeocystin), 4-phosphoryloxy-N,N,N-trimethyltryptamine (aeruginascin), 4-hydroxy-N-methyltryptamine (norpsilocin) and 4-phosphoryloxytryptamine (norbaeocystin) as well as β-carbolines such as harmine and harmaline, and terpenes ^14–18. Little is known about the effects of these additional components of psychedelic mushrooms in animal models, although there is accumulating evidence regarding the effect of some of the tryptamines of the psilocybin biosynthetic chain. Glatfelter et al ¹⁹ recently showed that only the tertiary amines, psilocybin, psilocin, and 4-acetoxy-N,N-dimethyltryptamine (psilacetin) induced HTR in mice while secondary amines such as baeocystin and norpsilocin and quaternary ammonium compounds such as aeruginascin had little or no effect. It is possible, however, that thesetryptamines present in psychedelic mushroom extracts may modulate the effects of psilocybin. In particular, it has been speculated that baeocystin or norpsilocin could potentially contribute to variable subjective effects ²⁰. Furthermore, additional components of mushroom extract such as beta-carbolines may exert biologically meaningful effects in spite of their low concentrations.

Recent research focusing on the possible therapeutic mechanisms of psychedelic agents has emphasized the putative role of sustained neuroplastic effects ^{21, 22}. These have been demonstrated by techniques that show an increase in dendritic spines and other microscopic measures of synaptic plasticity induced by psychedelics in cell cultures ²³ and in living mouse brain by single photon microscopy ²⁴. This therapeutic mechanism has received considerable emphasis in the development of novel, non-hallucinogenic, psychedelic agents ^25–27.

In this context, it is of considerable mechanistic and translational interest to determine whether PME differs from PSIL in its effects on HTR, induction of neuroplasticity and in the spectrum of metabolic effects that it induces. These questions were the focus of the current study. Recently, we comprehensively characterized HTR induced by PSIL in male C57Bl\6j mice ²⁸. Whether the characteristics of HTR induced by PME differ from those induced by PSIL under the same experimental conditions and at the same psilocybin dose, is an intriguing open question. A further key question is whether PSIL and PME differ in their effects on neuroplasticity which is increasingly regarded as playing a key role in the beneficial effects of psychedelics on depression and other psychiatric disorders ^{21, 22}. It is well established that synaptic proteins are pivotally involved in formation, elimination and pruning of synapses and modulation of their strength and plasticity ²⁹. Synaptic proteins are also involved in synaptic transmission and in maintaining synaptic structure and stability ³⁰. In accordance with this role, we studied the synaptic proteins GAP43, PSD95, synaptophysin and SV2A as markers of synaptic plasticity induced by PSIL and PME. Finally, we performed a metabolomic analysis of the effect of PSIL and PME on the mouse frontal cortex. Metabolomics is an emerging field that seeks to systematically characterize the unique chemical fingerprint that low-molecular-weight molecules leave behind after a given stimulus in a biological specimen ³¹. Alterations in biochemical processes such as energy production, oxidative stress and neurotransmission have been linked to various psychiatric disorders including MDD ³². A recent study analyzed the effects of ayahuasca treatments on primary astrocytes, revealing alterations in various biochemical pathways ³³. We set out to assess cortical metabolomic effects of PSIL and PME in order to identify key pathways that are involved in long-term sustained effects and to potentially identify metabolic pathways that are common to, or differentiate PSIL and PME.

Animals

Experiments were performed on adult (~ 11 weeks old) C57BL/6J male mice. Animals were housed under standardized conditions with a 12-h light/dark cycle, stable temperature (22 ± 1°C), controlled humidity (55 ± 10%) and free access to food and water. Experiments were conducted in accordance with AAALAC guidelines and were approved by the Authority for Biological and Biomedical Models Hebrew University of Jerusalem, Israel, Animal Care and Use Committee. All efforts were made to minimize animal suffering and the number of animals used.

Drugs

PSIL was supplied by Usona Institute, (Madison WI, USA) and was determined by AUC at 269.00 nm (UPLC) to contain 98.75 wt.% psilocybin. PME was supplied by Back of the Yards Algae Sciences - Parow Entheobiosciences (Chicago, IL, USA) and was determined to contain psilocybin 1.3 wt.%, psilocin 0.17 wt.%, norpsilocin 0.01 wt.%, baeocystin 0.02 wt.%, norbaeocystin 0.007 wt.% and aerugeniscin 0.008 wt.% by LC-MS/MS. PSIL and PME were dissolved in 0.9% NaCl. Whether administered PSIL or PME, mice received psilocybin at a dose of 4.4 mg/kg intraperitoneally (i.p.). For PME the dose was based on the following calculation: To prepare a solution for a mouse that weighs 30g so that the mouse will receive 4.4 mg/kg of pure psilocybin, \(30\times 0.44=0.132 \text{m}\text{g}\) of psilocybin per mouse is needed. Since 1mg of psilocybin is found in 76.92mg of PME (1.3%), \(76.92\times 0.132=10.15 \text{m}\text{g}\) of PME is needed per mouse. In all cases, injections were administered in a standard injection volume of 10 µl/g per mouse.

PME cultivation, extraction, preparation and analysis

Psilocybe cubensis was cultivated from genetically confirmed spore material (see www.Entheome.org for whole-genome sequence) germinated on light malt extract agar in a petri dish. Agar-cultured colonies were transferred to sterilized grain jars (rye, white millet, and oats). The colonized grain was transferred into pasteurized bulk substrate bags. The mushrooms were fruited and harvested after 18–28 days. The biomass was dried at 65°C for 6 hours. The dried ground biomass was extracted at a ratio of 1g of biomass to 20 mL of methanol for 24 hours at room temperature in stirred reaction vessels, and filtered through a Büchner funnel whereafter a second extraction took place for a further 24 hours. The filtrate from the second extraction was then combined with the filtrate from the first extraction and the methanol was removed using a rotary evaporator. The extract was then suspended in sterile water and spray dried using a nitrogen electrostatic spray dryer (FluidAir, Chicago, Illinois, USA) at conditions of inlet temperature − 120˚C, outlet temperature − 80˚C. A Waters Acquity H-Class UPLC and Xevo TQ-S Micro MS and Waters HSS T3 1.8 µm column (2.1 mm x 50 mm) (Waters Corporation, Milford, MA, USA) was used for determining tryptamine concentrations. A seven-point standard curve (10 ppb − 1 ppm) is used to calculate concentration from the detector response. It was produced using analytical standards produced by Cerilliant (Round Rock, TX, USA) (psilocybin and psilocin) and Usona (Madison, WI, USA) (norpsilocin, baeocystin, norbaeocystin, and aeruginascin).

Head Twitch Response

Head twitch response (HTR) was measured over 20 minutes by means of a magnetometer apparatus as described by de la Fuente Revenga et al. ³⁴ and Shahar et al. ²⁸. Briefly, small neodymium magnets (N50, 3mm diameter × 1mm height, 50 mg), were attached to the outer ears of mice. After a 5–7-day recovery period, the ear-tagged animals were placed inside a magnetometer apparatus (supplied by Mario de la Fuente Revenga PhD. of Virginia Commonwealth University) immediately after injection of vehicle, PSIL, or PME. The output was amplified (Pyle PP444 phono amplifier) and recorded at 1000 Hz using a NI USB-6001 (National Instruments, US) data acquisition system. Recordings were performed using a MATLAB driver (MathWorks, US, R2021a version, along with the NI myDAQ support package) with the corresponding National Instruments support package for further processing. A custom MATLAB script was used to record the processed signal, which was presented as graphs showing the change in current as peaks (mAh). A custom graphic user interface created in our laboratory was used to further process the recording into an Excel spreadsheet.

Western blotting

The frontal cortex, amygdala, striatum and hippocampus were dissected and stored at -80˚C, lysed in Pierce RIPA sample buffer (Thermo Scientific, USA), supplemented with protease inhibitor cocktail (Roche Diagnostics, Germany) and boiled for 10 min. Equivalent amounts of protein extracts (20 mg) were analyzed by SDS–12% PAGE, followed by transfer of the proteins to polyvinylidene fluoride membrane. Blots were blocked in 5% fat free milk in TBST buffer (Tris-Tween-buffered saline) and incubated in primary antibodies, one hour at room temperature. Primary antibodies included rabbit anti-GAP43 (ab75810, 1:2000; Abcam, UK), rabbit anti-PSD95 (ab238135, 1:2000; Abcam, UK), rabbit anti-Synaptophysin (ab32127, 1:2000; Abcam, UK), rabbit anti-SV2A (ab54351, 1:100; Abcam, UK) and mouse anti-β-Actin (8H10D10, 1:5000, Cell Signaling Technology). Blots were washed 3 times and incubated with a horseradish peroxidase-conjugated secondary antibodies (1:5000, ABclonal, China) for 1 h, followed by repeated washing with TBST buffer. Proteins were visualized by using enhanced chemiluminescence (ChemiDoc Reader MP, Bio-Rad, USA). The amount of each phosphorylated protein was normalized to the amount of the corresponding total protein detected in the sample and measured by intensity of β-actin.

Metabolomics

Brain tissue (frontal cortex) from mice administered PSIL, PME and VEH (n = 5 each), sacrificed 11 days later and stored at -80˚C was used for metabolomics assays performed by Human Metabolome Technologies-America (Boston, MA, USA). Brain tissue was placed in a homogenization tube, along with zirconia beads and a solution containing 50% acetonitrile in Milli-Q water (v/v) containing HMT internal standards, after which the sample was homogenized (using beads shaker) at 4˚C for 120 sec. The homogenate was then centrifuged at 2,300 × g, 4˚C for 5 min. The upper aqueous layer was centrifugally filtered at 4˚C through a 5-kDa cut-off filter. The filtrate was evaporated to dryness under vacuum and reconstituted in Milli-Q water for metabolome analysis. Metabolomic analysis was performed for PSIL, PME and VEH treated mice using untargeted CE-FTMS ³⁵. Extracted metabolites were measured in the Cation and Anion modes of CE-FTMS based metabolome analysis. An Agilent CE system and Q Exactive Plus mass spectrometer were used for both anionic and cationic measurements. The CE capillary was an uncoated fused silica capillary i.d. 50 µm × 80 cm using HMT running and sheath buffers with mass scan range of m/z 70 to 1,050. MetaboAnalyst 5.0 software was used to analyze the data and identify possible associated metabolic pathways and treatment-differentiated metabolic expression patterns.

Statistical analysis

The experimental data are expressed as the mean ± standard error of the mean (SEM). To determine inter group differences, one- and two-way repeated measure analysis of variance (ANOVA) were used as indicated. Tukey’s or Dunnett’s Multiple Comparison Tests were used to analyze post-hoc comparisons. p < 0.05 (two tailed) was the criterion for significance. Graph Pad Prism, version 9.3.1 software was used for all statistical analyses.

Head Twitch Response

We assessed the effect of PSIL and PME on mouse HTR at a psilocybin dose of 4.4 mg/kg as a representative human clinical trial dose as previously described ²⁸ (Fig. 1). HTR over time (2 min time bins over 20 minutes) showed a strong increase of HTR (Fig. 1a) compared to vehicle reflected in a between-subject main effect of vehicle vs PSIL/PME in a two-way repeated measure ANOVA (Time x Treatment F [18, 333] = 10.86 p < 0.0001, Treatment F [2, 37] = 52.18 p < 0.0001), and no difference was observed between PSIL and PME reflected in a within-subject main effect of PSIL vs PME in a two-way repeated measure of ANOVA (Time x Treatment F [9, 252] = 1.162 p = 0.3197, Treatment F [1, 28] = 0.04689 p = 0.8301). Total and peak PSIL/PME-induced HTR during the 20 min measuring period were compiled (Fig. 1b, c). One-way ANOVA and post hoc Šídák's analysis of total and peak HTR show a significant difference that both treatments compared to vehicle, with no differences between them (Total HTR: Vehicle vs PSIL/PME p < 0.0001, PSIL vs PME p = 0.9419. Peak HTR: Vehicle vs PSIL/PME p < 0.0001, PSIL vs PME p = 0.8662.) These HTR results are for the same mice that participated in the synaptic protein assays. Supplementary Fig. S1 shows HTR results for a larger sample of mice (including those reported above) that were tested for HTR. Similarly, to the previous results, no difference between PSIL and PME was observed.

Synaptic Proteins

Using Western blot analysis, we quantified 4 synaptic proteins (GAP43, PSD95, synaptophysin and SV2A – Figs. 2–5) in 4 brain areas (frontal cortex, hippocampus, amygdala and striatum) 11 days after intraperitoneal injection of PSIL and PME, both at 4.4 mg/kg psilocybin. Initially we analyzed each synaptic protein in each brain area separately. Our key findings were:

(1) PSIL and PME significantly increased GAP43 levels in the frontal cortex (PSIL p = 0.019; PME p = 0.039) and hippocampus (PSIL p = 0.015; PME p = 0.027).

(2) PSIL and PME significantly increased synaptophysin in the hippocampus (PSIL, p = 0.041; PME, p = 0.05) and amygdala (PSIL, p = 0.035; PME, p = 0.004). In the striatum, synaptophysin was increased by PME only (p = 0.023).

(3) PSIL increased SV2A in the amygdala (p = 0.036) and PME did so in the hippocampus (p = 0.014) (Fig. 5).

(4) There were no significant effects of PSIL or PME on PSD95 in any brain area when these were analyzed separately.

Supplementary Figs. S2-5 show effects of PSIL and PME on synaptic proteins in each of the 4 brain areas separately.

(1) Hippocampus was the brain area in which the greatest number of significant effects of PSIL and PME were observed. PSIL and PME both significantly increased GAP43 (p = 0.015; p = 0.027, respectively) and synaptophysin (p = 0.041; p = 0.035, respectively). PME (p = 0.014) but not psilocybin increased SV2A levels.

(2) In the amygdala, synaptophysin was increased by PSIL (p = 0.035) and PME (p = 0.004) and SV2A by PSIL (p = 0.036) but not PME.

(3) In the frontal cortex GAP43 was significantly increased by both PSIL (p = 0.018) and PME (p = 0.039).

(4) In the striatum, synaptophysin was increased by PME only (p = 0.026).

A stronger effect of PME on synaptic proteins was highlighted by comparing overall effects of PSIL and PME across all 4 brain areas. We performed a nested ANOVA for each synaptic protein separately (Fig. 6a-d). The results showed a significant overall increase in GAP43 (F [2, 223] = 6.354, p = 0.002). PME (p = 0.0009) but not PSIL was significant versus vehicle on post hoc testing (Fig. 6a). For PSD95 there was also a significant overall increase across all brain areas (F [2, 55] = 4.599, p = 0.014). PSIL (p = 0.01) and PME (p = 0.022) were both significant versus vehicle on post hoc testing (Fig. 6b). Synaptophysin was also significantly increased across all 4 brain areas (F [2, 56] = 8.563, p = 0.0006). On post-hoc testing only PME was significant (p = 0.0002) (Fig. 6c). For SV2A the overall effect of treatment was significant across all brain areas (F [2, 113] = 4.21, p = 0.017) and both PSIL (p = 0.019) and PME (p = 0.033) were significant on post-hoc testing (Fig. 6d). Thus, when overall effects across all four brain regions were taken into account, PME increased all 4 synaptic proteins while PSIL increased only 2 of them.

We performed a second nested ANOVA in which effects of PSIL and PME on all 4 synaptic proteins were examined in each brain area separately (Supplementary Fig. S6a-d). A stronger effect of PME was evident in 3 out of 4 brain areas. In the frontal cortex there was a significant overall effect of treatment (F [2, 57] = 3.579, p = 0.034). PME (p = 0.028), but not PSIL, was significant versus vehicle on overall testing (Fig. S6a). There was an overall effect of treatment in the hippocampus too (F [2, 58] = 6.083, p = 0.004). In this case both PSIL (p = 0.005) and PME (p = 0.011) were significant (Fig. S6c). In the amygdala (F [2, 56] = 5.273, p = 0.008) (Fig. S6b) and striatum (F [2, 55] = 3.516, p = 0.034) (Fig. S6d) there was an overall effect of treatment but only PME (p = 0.007; p = 0.023, respectively) was significant on post-hoc testing.

To determine whether effects on synaptic proteins might be discernable after PSIL and PME administration at an earlier stage than 11 days, we examined levels of three synaptic proteins (GAP43, PSD95 and synaptophysin) in 4 brain areas 3 days after treatment administration (Supplementary Figs. S7-10). There were significant effects of PSIL (p = 0.010) and PME (p = 0.037) to increase GAP43 in the striatum. No effects on GAP43 in frontal cortex, hippocampus and amygdala were significant nor effects on PSD95 and synaptophysin in any of the 4 brain areas.

We further examined whether there was any relationship between the HTR induced by PSIL and PME immediately after injection and the effect of these agents on synaptic proteins 11 days later. We compared both total HTR and peak HTR that each compound induced with each of the 4 synaptic proteins in each of the 4 brain areas totaling a total of 64 simple linear regression correlation tests. There was no correlation between synaptic protein increase to any HTR measurement (total or peak HTR) (Supplementary Figs. S11-27).

Metabolomics

The CE-FTMS polar method enabled detection of 466 library annotated metabolites, 230 quantitated metabolites and an additional 1566 features with partial identification. Partial least squares-discriminant analysis with MetaboAnalyst 5.0, was used to compare the metabolomic profiles of PME, PSIL, VEH (Fig. 7a). The results showed clear separation between PME and VEH but not between PSIL and vehicle nor between PSIL and PME. The principal metabolites contributing to this separation were 3-phenyllactic acid and cystine (under-expressed in the PME group) and phosphoenolpyruvic acid (PEP) (over-expressed in the PME group) (Fig. 7b). In addition, the purines guanosine, hypoxanthine and inosine showed a progressive decline from vehicle to PSIL to PME groups, while their phosphorylated forms are progressively upregulated (Supplementary Fig. S27). Statistical analysis comparing the fold change (1.25 cutoff) between PME vs. VEH revealed many differentiated metabolites (Supplementary Fig. S28), where guanosine and 6-aminohexanoic acid were found to be significant, withstanding FDR correction (FDR < 0.1). A heatmap visualizing specific pathway analysis, based on the musculus (KEGG) pathway library, between PME and VEH is presented in Fig. 8. The following pathways were found to be significantly differentiated between the groups (Table 1): Purine, pyrimidine, arginine and proline, tryptophan and beta-alanine metabolism.

Table 1

Significantly differentiated metabolic pathways between PME and VEH.
Pathway	Metabolites	Total	Expected	Hits	p Value	FDR
Purine metabolism	Guanosine Inosine Hypoxanthine Xanthine Xanthosine AMP	66	0.705	6	3.30E-05	0.00277
Pyrimidine metabolisms	5-Oxoproline Putrescine Cadaverine Spermine	28	0.299	4	0.000153	0.00641
Arginine and proline metabolism	Putrescine N-Acetylputrescine Spermine Phosophocreatine	38	0.406	4	0.000516	0.0144
Tryptophan metabolism	Spermine Uracil	21	0.224	2	0.02	0.419
beta-Alanine metabolism	Riboflavin	4	0.0427	1	0.0421	0.707

In a comprehensive series of studies that span immediate effects on HTR, changes in synaptic protein levels 3 and 11 days after treatment administration and changes in metabolic parameters at 11 days, we compared the effects of PSIL and PME at the equivalent psilocybin dose. Our findings show no difference in acute effects on HTR. However, we found an effect of PME on synaptic protein levels in 4 brain areas that is significantly more pronounced overall than the effect of PSIL. At the same time point (11 days after treatment administration), the effect of PME on metabolic parameters in the frontal cortex is clearly defined from that of PSIL suggesting a discernibly different or quantitatively stronger therapeutic mechanism.

Our HTR findings contrast with those of Zhuk et al ¹² who compared the effect of psilocin, the active metabolite of psilocybin, on HTR to those of two different psilocin-containing mushroom extracts, Ph. cyanopus and P. semilanceata. Zhuk et al ¹² found that the two mushroom extracts induced a similar number of HTR compared to pure psilocin, despite the fact that the amount of psilocin present in the extracts was much lower. Zhuk et al ¹² speculated that this phenomenon could be related to the synergistic effect of the other indole alkaloids in the extracts. An important difference between the two studies is that our comparison was based on psilocybin and not psilocin content and the PME and PSIL that we injected were held constant in terms of psilocybin content. Thus, the two studies are not directly comparable.

Our second key focus was on comparing effects of PME and PSIL on key synaptic proteins, that we studied as markers of neuroplasticity. Neuroplasticity refers to the lifelong capacity of the brain to respond to experiences, learning and the environment and to reorganize structure, function and connections in response to such stimuli. Neuroplastic effects may be defined and measured in structural and functional terms. With regard to the effect of psychedelics, a key focus in the domain of neuroplastic effects has been on structural changes in the synapse with emphasis on growth of new dendrites and proliferation of dendritic spines ^{1, 23, 24, 36}. It is widely suggested that synaptic plasticity is the central mechanism whereby psychedelics achieve their therapeutic effects ^{21, 22, 37–39}.

Synaptic proteins are pivotally involved in synaptic plasticity by regulating synaptic transmission and because of their role in structural changes that occur during plasticity ⁴⁰. Synaptic proteins such as GAP43, PSD95, synaptophysin and SV2A play vital roles in the development, functioning, and plasticity of the nervous system ²⁹. GAP-43 plays a crucial role in axonal outgrowth, guidance, and pathfinding during development ^{41, 42}. It has been implicated in the remodeling of synaptic connections, the stabilization of newly formed synapses, and the modulation of neurotransmitter release. Studies have shown that changes in the expression and localization of GAP-43 are associated with synaptic plasticity and learning processes ^{43, 44}. PSD-95 interacts with various binding partners, such as associated synaptic proteins, ion channels, and cytoskeletal elements, to form a complex network that stabilizes and regulates synaptic function, transmission and formation and maintenance of dendritic spines, receptor trafficking, and synaptic signaling and plasticity mechanisms ^45–48. Synaptophysin is involved in synaptic plasticity ⁴⁹, and in synaptic vesicle exocytosis ^{50, 51}. It is an integral component of the synaptic vesicle membrane and interacts with other proteins to regulate vesicle fusion with the presynaptic membrane, thereby facilitating the release of neurotransmitters into the synapse ^52–54. SV2A is involved in multiple aspects of synaptic vesicle function and neurotransmitter release ⁵⁵. It plays a crucial role in regulating the trafficking and exocytosis of synaptic vesicles ^{46, 56}. The absence of SV2A leads to a decrease in neurotransmission mediated by action potentials, while neurotransmission independent of action potentials remains unaffected ^{56, 57}.

Prior evidence that psilocybin or any other psychedelic drugs influence brain levels of the synaptic proteins that we studied, is limited to one report. Raval et al. ⁵⁸ examined the density of SV2A in pig brain one day and 7 days after a 0.08mg/kg of psilocybin. SV2A protein density was determined by [3H]UCB-J autoradiography in the hippocampus and the prefrontal cortex. Compared to the saline-treated group, psilocybin treatment was associated with 4.42% higher SV2A in the hippocampus (p < 0.0001), one day after psilocybin injection and 9.24% higher SV2A in the hippocampus (p = 0.024), seven days after psilocybin. Our results partially overlap with those of Raval et al. ⁵⁸. We found that in mice, a dose of PME containing a similar psilocybin dose (adjusted for species differences), increased SV2A in the hippocampus 11 days after administration and a similar dose of PSIL increased SV2A in the striatum, an area that Raval et al. ⁵⁸ did not examine.

Taking into account both PME and PSIL, it is noteworthy that we found an overall increase in all 4 synaptic proteins examined (GAP43, PSD95, synaptophysin and SV2A) over all 4 brain areas (frontal cortex, hippocampus, amygdala, striatum). The stronger effect of PME was highlighted by the fact the extract increased all four synaptic proteins, while PSIL increased only two of them. These findings suggest that synaptic protein levels can serve as an informative marker for the effect of psychedelic compounds on synaptic plasticity. Further studies are required to understand regional differences in the effect of PME and PSIL on the synaptic proteins that we studied. It will also be of considerable interest to compare the effect of PME and PSIL on brain derived neurotrophic factor (BDNF) levels. Given the cardinal role of BDNF in supporting growth, survival, and differentiation of both developing and mature neurons and inducing synaptic plasticity, its relationship to psychedelic compounds has been extensively tested in the past years. Animal models have established that BDNF expression is elevated after a single administration of psychedelics ⁵⁹, resulting in changed neuroplasticity including dendritic complexity, which outlasted the acute effects of the psychedelic. Moreover, chronic psychedelic administrations increased BDNF mRNA and protein levels up to a month after treatment ^60–62. Human studies have usually assessed peripheral plasma BDNF protein levels as a biomarker for neuroplastic modulation, although results are mixed ²². Most recently, psychedelics have been found to directly bind to the BDNF receptor TrKB, but the therapeutic implications of this interaction are not fully understood ⁶³.

In our metabolomics analyses, principal components analysis revealed complete separation between PME and VEH groups, whereas PSIL components corresponded to a shared area associated with both groups. This initial finding is expected given the natural complexity of biological specimens, as compared to a single chemical molecule, when compared to VEH and may explain the progressive differences in specific metabolite expression. In this context we propose a “purine hypothesis” where “free” purines in their unphosphorylated form show a progressive decline from VEH, to PSIL, to PME while the phosphorylated (bioactive) forms are progressively up-regulated in the counter direction. This may in part explain the energy-dependent biological processes that are attributed to the potentially therapeutic effects of PSIL and PME (anti-inflammatory, neurite outgrowth, neurogenesis, neuroprotection and more) ⁶⁴. This system level observation may further demonstrate key longitudinal biotransformation changes (PEP associated ATP and neurotransmitter synthesis, synaptic vesicle formation etc.) that may underlie additional energy supply that enables positive neuroplastic modulations ⁶⁵. Furthermore, signaling pathway analysis supported this hypothesis, where the purine metabolism pathway was found to be the most significantly differentiated between PME and VEH. Other metabolic pathways that were found to be significantly different were the pyrimidine metabolism pathway associated with nucleic acid synthesis, and the arginine and proline metabolism pathway associated with metabolism of several amino acids including arginine, ornithine, proline, citrulline, and glutamate in mammals.

In conclusion, we have shown, in a mouse model, clear synaptic protein pattern differences following the administration of PME and PSI and have demonstrated for the first-time, alterations in cortical metabolic expression patterns between PME and vehicle treated mice and have suggested possible pathway-related mechanisms to explain these differences. While our data do not provide conclusive evidence for the therapeutic superiority of naturally-derived psychedelic mushroom extract over chemical psilocybin, they open the door to serious consideration of the potential of combinations of molecules found in psychedelic mushrooms, especially those related to the psilocybin biosynthetic pathway, with psilocybin. Such combinations may not only have enhanced or more prolonged therapeutic effects but may result in even more effective combinations by increasing the amount of the additional neuroactive compounds that are only present in extremely small amounts naturally.

Acknowledgments

Masha Chaykin expertly assisted with Western Blot assays.

Support

The Hadassah BrainLabs Center for Psychedelic Research was founded with the support of Negev Labs

This work was supported in part by Back of the Yards algae sciences (BYAS) and Parow Entheobiosciences (ParowBio)

Conflict of Interest

Leonard Lerer is the Founder and CEO of Back of the Yards algae sciences (BYAS) and a Founder of Parow Entheobiosciences (ParowBio).

Karin Blakolmer is a Founder and CEO of Parow Entheobiosciences (ParowBio).

Bernard Lerer is a consultant to Back of the Yards algae sciences (BYAS) and Parow Entheobiosciences (ParowBio).

A patent application has been submitted by ParowBio based on the work reported here.

Alex Buko is an employee of Human Metabolomics Technologies

McClure-Begley TD, Roth BL. The promises and perils of psychedelic pharmacology for psychiatry. Nature Reviews Drug Discovery 2022; 21(6): 463-473.
Strauss D, Ghosh S, Murray Z, Gryzenhout M. An Overview on the Taxonomy, Phylogenetics and Ecology of the Psychedelic Genera Psilocybe, Panaeolus, Pluteus and Gymnopilus. Frontiers in Forests and Global Change 2022; 5.
Barker SA. N, N-Dimethyltryptamine (DMT), an Endogenous Hallucinogen: Past, Present, and Future Research to Determine Its Role and Function. Front Neurosci 2018; 12: 536.
Uthaug MV, Lancelotta R, van Oorsouw K, Kuypers KPC, Mason N, Rak J et al. A single inhalation of vapor from dried toad secretion containing 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT) in a naturalistic setting is related to sustained enhancement of satisfaction with life, mindfulness-related capacities, and a decrement of psychopathological symptoms. Psychopharmacology 2019; 236(9): 2653-2666.
Dinis-Oliveira RJ, Pereira CL, da Silva DD. Pharmacokinetic and Pharmacodynamic Aspects of Peyote and Mescaline: Clinical and Forensic Repercussions. Curr Mol Pharmacol 2019; 12(3): 184-194.
Schiff PL. Ergot and its alkaloids. Am J Pharm Educ 2006; 70(5): 98.
Ferber SG, Namdar D, Hen-Shoval D, Eger G, Koltai H, Shoval G et al. The "Entourage Effect": Terpenes Coupled with Cannabinoids for the Treatment of Mood Disorders and Anxiety Disorders. Curr Neuropharmacol 2020; 18(2): 87-96.
Jikoms N. Do magic mushrooms have ‘strains’ like cannabis? 2022. https://www.leafly.com/news/science-tech/do-magic-mushrooms-have-strains-like-cannabis. Accessed June 27, 2023.
FreshCap Mushrooms. Do All Magic Mushrooms Have The Same Effect? YouTube. 2023. https://www.youtube.com/watch?v=PBEHe5cC9qg. Accessed June 27, 2023.
Gartz J. Analysis of Aeruginascin in Fruit Bodies of the Mushroom Inocybe aeruginascens. International Journal of Crude Drug Research 1989; 27(3): 141-144.
Matsushima Y, Shirota O, Kikura-Hanajiri R, Goda Y, Eguchi F. Effects of Psilocybe argentipes on marble-burying behavior in mice. Bioscience, biotechnology, and biochemistry 2009; 73(8): 1866-1868.
Zhuk O, Jasicka-Misiak I, Poliwoda A, Kazakova A, Godovan VV, Halama M et al. Research on acute toxicity and the behavioral effects of methanolic extract from psilocybin mushrooms and psilocin in mice. Toxins 2015; 7(4): 1018-1029.
Van Court RC, Wiseman MS, Meyer KW, Ballhorn DJ, Amses KR, Slot JC et al. Diversity, biology, and history of psilocybin-containing fungi: Suggestions for research and technological development. Fungal Biology 2022; 126(4): 308-319.
Blei F, Dörner S, Fricke J, Baldeweg F, Trottmann F, Komor A et al. Simultaneous Production of Psilocybin and a Cocktail of β-Carboline Monoamine Oxidase Inhibitors in “Magic” Mushrooms. Chemistry – A European Journal 2020; 26(3): 729-734.
Leung AY, Paul AG. Baeocystin and norbaeocystin: new analogs of psilocybin from Psilocybe baeocystis. J Pharm Sci 1968; 57(10): 1667-1671.
Gotvaldová K, Hájková K, Borovička J, Jurok R, Cihlářová P, Kuchař M. Stability of psilocybin and its four analogs in the biomass of the psychotropic mushroom Psilocybe cubensis. Drug Testing and Analysis 2021; 13(2): 439-446.
Lenz C, Wick J, Hoffmeister D. Identification of ω-N-Methyl-4-hydroxytryptamine (Norpsilocin) as a Psilocybe Natural Product. Journal of Natural Products 2017; 80(10): 2835-2838.
Dörner S, Rogge K, Fricke J, Schäfer T, Wurlitzer JM, Gressler M et al. Genetic Survey of Psilocybe Natural Products. ChemBioChem 2022; 23(14): e202200249.
Glatfelter GC, Pottie E, Partilla JS, Sherwood AM, Kaylo K, Pham DNK et al. Structure–Activity Relationships for Psilocybin, Baeocystin, Aeruginascin, and Related Analogues to Produce Pharmacological Effects in Mice. ACS Pharmacology & Translational Science 2022; 5(11): 1181-1196.
Sherwood AM, Halberstadt AL, Klein AK, McCorvy JD, Kaylo KW, Kargbo RB et al. Synthesis and biological evaluation of tryptamines found in hallucinogenic mushrooms: norbaeocystin, baeocystin, norpsilocin, and aeruginascin. Journal of natural products 2020; 83(2): 461-467.
Grieco SF, Castrén E, Knudsen GM, Kwan AC, Olson DE, Zuo Y et al. Psychedelics and Neural Plasticity: Therapeutic Implications. The Journal of Neuroscience 2022; 42(45): 8439-8449.
Calder AE, Hasler G. Towards an understanding of psychedelic-induced neuroplasticity. Neuropsychopharmacology 2023; 48(1): 104-112.
Ly C, Greb AC, Cameron LP, Wong JM, Barragan EV, Wilson PC et al. Psychedelics Promote Structural and Functional Neural Plasticity. Cell Rep 2018; 23(11): 3170-3182.
Shao L-X, Liao C, Gregg I, Davoudian PA, Savalia NK, Delagarza K et al. Psilocybin induces rapid and persistent growth of dendritic spines in frontal cortex in vivo. bioRxiv 2021: 2021.2002.2017.431629.
Cao D, Yu J, Wang H, Luo Z, Liu X, He L et al. Structure-based discovery of nonhallucinogenic psychedelic analogs. Science 2022; 375(6579): 403-411.
Cameron LP, Tombari RJ, Lu J, Pell AJ, Hurley ZQ, Ehinger Y et al. A non-hallucinogenic psychedelic analogue with therapeutic potential. Nature 2021; 589(7842): 474-479.
Kaplan AL, Confair DN, Kim K, Barros-Álvarez X, Rodriguiz RM, Yang Y et al. Bespoke library docking for 5-HT(2A) receptor agonists with antidepressant activity. Nature 2022; 610(7932): 582-591.
Shahar O, Botvinnik A, Esh-Zuntz N, Brownstien M, Wolf R, Lotan A et al. Role of 5-HT2A, 5-HT2C, 5-HT1A and TAAR1 receptors in the head twitch response induced by 5-hydroxytryptophan and psilocybin: Translational implications. International Journal of Molecular Sciences 2022; 23(22): 14148.
Nazir FH, Becker B, Brinkmalm A, Höglund K, Sandelius Å, Bergström P et al. Expression and secretion of synaptic proteins during stem cell differentiation to cortical neurons. Neurochemistry International 2018; 121: 38-49.
Truckenbrodt S, Viplav A, Jähne S, Vogts A, Denker A, Wildhagen H et al. Newly produced synaptic vesicle proteins are preferentially used in synaptic transmission. The EMBO Journal 2018; 37(15): e98044.
Courant F, Antignac J-P, Dervilly-Pinel G, Le Bizec B. Basics of mass spectrometry based metabolomics. PROTEOMICS 2014; 14(21-22): 2369-2388.
Mandal PK, Gaur S, Roy RG, Samkaria A, Ingole R, Goel A. Schizophrenia, Bipolar and Major Depressive Disorders: Overview of Clinical Features, Neurotransmitter Alterations, Pharmacological Interventions, and Impact of Oxidative Stress in the Disease Process. ACS Chemical Neuroscience 2022; 13(19): 2784-2802.
Zandonadi FS, Silva AAR, Melo AA, Ignarro RS, Matos TS, Santos EA et al. Understanding ayahuasca effects in major depressive disorder treatment through in vitro metabolomics and bioinformatics. Analytical and Bioanalytical Chemistry 2023: 1-18.
de la Fuente Revenga M, Vohra HZ, González-Maeso J. Automated quantification of head-twitch response in mice via ear tag reporter coupled with biphasic detection. Journal of neuroscience methods 2020; 334: 108595.
Sugishita T, Tokunaga M, Kami K, Terai K, Yamamoto H, Shinohara H et al. Determination of the Minimum Sample Amount for Capillary Electrophoresis-Fourier Transform Mass Spectrometry (CE-FTMS)-Based Metabolomics of Colorectal Cancer Biopsies. Biomedicines 2023; 11(6): 1706.
Jones KA, Srivastava DP, Allen JA, Strachan RT, Roth BL, Penzes P. Rapid modulation of spine morphology by the 5-HT2A serotonin receptor through kalirin-7 signaling. Proceedings of the National Academy of Sciences 2009; 106(46): 19575-19580.
Slocum ST, DiBerto JF, Roth BL. Molecular insights into psychedelic drug action. Journal of Neurochemistry 2022; 162(1): 24-38.
de la Fuente Revenga M, Zhu B, Guevara CA, Naler LB, Saunders JM, Zhou Z et al. Prolonged epigenomic and synaptic plasticity alterations following single exposure to a psychedelic in mice. Cell Reports 2021; 37(3): 109836.
Lukasiewicz K, Baker JJ, Zuo Y, Lu J. Serotonergic Psychedelics in Neural Plasticity. Frontiers in Molecular Neuroscience 2021; 14.
Lohmann C, Kessels HW. The developmental stages of synaptic plasticity. J Physiol 2014; 592(1): 13-31.
Console-Bram LM, Fitzpatrick-McElligott SG, McElligott JG. Distribution of GAP-43 mRNA in the immature and adult cerebellum: a role for GAP-43 in cerebellar development and neuroplasticity. Developmental brain research 1996; 95(1): 97-106.
Cantallops I, Routtenberg A. Activity‐dependent regulation of axonal growth: Posttranscriptional control of the GAP‐43 gene by the NMDA receptor in developing hippocampus. Journal of neurobiology 1999; 41(2): 208-220.
Snipes G, Chan S, McGuire C, Costello B, Norden J, Freeman J et al. Evidence for the coidentification of GAP-43, a growth-associated protein, and F1, a plasticity-associated protein. Journal of Neuroscience 1987; 7(12): 4066-4075.
He Q, Dent EW, Meiri KF. Modulation of actin filament behavior by GAP-43 (neuromodulin) is dependent on the phosphorylation status of serine 41, the protein kinase C site. Journal of Neuroscience 1997; 17(10): 3515-3524.
Chen X, Levy JM, Hou A, Winters C, Azzam R, Sousa AA et al. PSD-95 family MAGUKs are essential for anchoring AMPA and NMDA receptor complexes at the postsynaptic density. Proceedings of the National Academy of Sciences 2015; 112(50): E6983-E6992.
Xu T, Bajjalieh SM. SV2 modulates the size of the readily releasable pool of secretory vesicles. Nature cell biology 2001; 3(8): 691-698.
Dore K, Malinow R. Elevated PSD-95 blocks ion-flux independent LTD: a potential new role for PSD-95 in synaptic plasticity. Neuroscience 2021; 456: 43-49.
Coley AA, Gao W-J. PSD95: A synaptic protein implicated in schizophrenia or autism? Progress in Neuro-Psychopharmacology and Biological Psychiatry 2018; 82: 187-194.
Gylys KH, Fein JA, Yang F, Wiley DJ, Miller CA, Cole GM. Synaptic changes in Alzheimer's disease: increased amyloid-β and gliosis in surviving terminals is accompanied by decreased PSD-95 fluorescence. The American journal of pathology 2004; 165(5): 1809-1817.
Shibaguchi H, Takemura K, Kan S, Kataoka Y, Kaibara M, Saito N et al. Role of synaptophysin in exocytotic release of dopamine from Xenopus oocytes injected with rat brain mRNA. Cellular and Molecular Neurobiology 2000; 20(3): 401-408.
Valtorta F, Pennuto M, Bonanomi D, Benfenati F. Synaptophysin: leading actor or walk‐on role in synaptic vesicle exocytosis? Bioessays 2004; 26(4): 445-453.
Alder J, Kanki H, Valtorta F, Greengard P, Poo MM. Overexpression of synaptophysin enhances neurotransmitter secretion at Xenopus neuromuscular synapses. Journal of Neuroscience 1995; 15(1): 511-519.
Daly C, Ziff EB. Ca2+-dependent formation of a dynamin-synaptophysin complex: potential role in synaptic vesicle endocytosis. Journal of Biological Chemistry 2002; 277(11): 9010-9015.
Horikawa HP, Kneussel M, El Far O, Betz H. Interaction of synaptophysin with the AP-1 adaptor protein γ-adaptin. Molecular and Cellular Neuroscience 2002; 21(3): 454-462.
Bartholome O, Van den Ackerveken P, Sánchez Gil J, de la Brassinne Bonardeaux O, Leprince P, Franzen R et al. Puzzling out synaptic vesicle 2 family members functions. Frontiers in molecular neuroscience 2017; 10: 148.
Crowder KM, Gunther JM, Jones TA, Hale BD, Zhang HZ, Peterson MR et al. Abnormal neurotransmission in mice lacking synaptic vesicle protein 2A (SV2A). Proceedings of the National Academy of Sciences 1999; 96(26): 15268-15273.
Janz R, Goda Y, Geppert M, Missler M, Südhof TC. SV2A and SV2B function as redundant Ca2+ regulators in neurotransmitter release. Neuron 1999; 24(4): 1003-1016.
Raval NR, Johansen A, Donovan LL, Ros NF, Ozenne B, Hansen HD et al. A Single Dose of Psilocybin Increases Synaptic Density and Decreases 5-HT2A Receptor Density in the Pig Brain. International Journal of Molecular Sciences 2021; 22(2): 835.
Nardai S, László M, Szabó A, Alpár A, Hanics J, Zahola P et al. N, N-dimethyltryptamine reduces infarct size and improves functional recovery following transient focal brain ischemia in rats. Experimental Neurology 2020; 327: 113245.
De Vos CM, Mason NL, Kuypers KP. Psychedelics and neuroplasticity: a systematic review unraveling the biological underpinnings of psychedelics. Frontiers in psychiatry 2021; 12: 724606.
Martin DA, Marona-Lewicka D, Nichols DE, Nichols CD. Chronic LSD alters gene expression profiles in the mPFC relevant to schizophrenia. Neuropharmacology 2014; 83: 1-8.
Colaço CS, Alves SS, Nolli LM, Pinheiro WO, de Oliveira DGR, Santos BWL et al. Toxicity of ayahuasca after 28 days daily exposure and effects on monoamines and brain-derived neurotrophic factor (BDNF) in brain of Wistar rats. Metabolic Brain Disease 2020; 35: 739-751.
Moliner R, Girych M, Brunello CA, Kovaleva V, Biojone C, Enkavi G et al. Psychedelics promote plasticity by directly binding to BDNF receptor TrkB. Nature Neuroscience 2023; 26(6): 1032-1041.
Kozlowska U, Nichols C, Wiatr K, Figiel M. From psychiatry to neurology: Psychedelics as prospective therapeutics for neurodegenerative disorders. Journal of Neurochemistry 2022; 162(1): 89-108.
Ishida A, Noda Y, Ueda T. Synaptic vesicle-bound pyruvate kinase can support vesicular glutamate uptake. Neurochemical research 2009; 34: 807-818.

The Hadassah BrainLabs Center for Psychedelic Research was founded with the support of Negev Labs This work was supported in part by Back of the Yards algae sciences (BYAS) and Parow Entheobiosciences (ParowBio) Leonard Lerer is the Founder and CEO of Back of the Yards algae sciences (BYAS) and a Founder of Parow Entheobiosciences (ParowBio). Karin Blakolmer is a Founder and CEO of Parow Entheobiosciences (ParowBio). Bernard Lerer is a consultant to Back of the Yards algae sciences (BYAS) and Parow Entheobiosciences (ParowBio). A patent application has been submitted by ParowBio based on the work reported here. Alex Buko is an employee of Human Metabolomics Technologies

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Distinctive Molecular and Metabolic Profiles of Chemically Synthesized Psilocybin and Psychedelic Mushroom Extract

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PME cultivation, extraction, preparation and analysis

Head Twitch Response

Western blotting

Metabolomics

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Head Twitch Response

Synaptic Proteins

Metabolomics

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