Designer Drugs Research Articles

Novel Psychoactive Substance Use and Psychological Trauma: A Multimethodological Analysis

Background Authors discuss the connections between novel psychoactive substance (NPS) use and psychological trauma. The transition from classical substances to NPS, a paradigm change, poses a challenge for the treatment systems. Objective: Research evidence suggests difficulties in emotion regulation and trauma-related NPS-use. Authors explore some demographic and psychopathological characteristics related to such findings and examine the connections between emotion regulation deficiency and the choice of substance. Method This study uses a methodological triangulation of a biologically identified sample to confirm NPS use, a survey method to describe users’ socioeconomic characteristics, and Minnesota Multiphasic Personality Inventory (MMPI-2) subscales to study dysfunctions in emotion regulation. Results Participants (77 patients) were mainly polydrug users. The transgenerational transfer of substance use was a salient feature, but material deprivation was not characteristic of the entire sample. NPS use was not connected to certain psychopathological characteristics the way classical substance use was. More than half of the respondents had elevated scores on MMPI-2 Demoralization (RCd) and Dysfunctional Negative Emotions (RC7) scales. Nearly half of them also scored high on Neuroticism/Negative Emotionality (NEGE). Conclusions Results suggest that NPS use in the context of polydrug use is connected to psychological trauma and emotion regulation deficiency, but the MMPI-2 scales to assess emotional dysfunctions are not connected to a particular type of NPS.

Rethinking Mental Automatism: De Clérambault's Theory in the Age of Novel Psychoactive Drugs: Psychotropic Effects and Synthetic Psychosis.

The widespread use of novel psychoactive substances (NPSs)-defined as new narcotic or psychotropic agents not classified under the Single Convention on Narcotic Drugs of 1961 or the Convention on Psychotropic Substances of 1971-poses a significant challenge to contemporary mental health paradigms due to their impact on psychiatric disorders. This study revisits and expands upon the theory of mental automatism as proposed by Gaëtan Gatian de Clérambault, aiming to elucidate the psychopathological mechanisms underlying substance-induced psychoses (SIP) and their distinction from non-induced psychoses (schizophrenia and related disorders). Through a phenomenological and clinical investigation, we explore the relevance of mental automatism in the development of toxic psychoses, drawing upon the historical and contemporary literature. This research highlights the psychopathological distinctions between induced and non-induced psychoses and the transition mechanisms from acute to chronic psychosis states. De Clérambault's theory, supplemented by Janet, Jackson, and Bonhoeffer's contributions, provides a foundational framework for understanding the genesis of SIP. Our findings suggest that NPS consumption, particularly among adolescents and psychiatric patients, significantly correlates with increased risks of SIP, marked by a transition to chronicity influenced by biological lesions triggered by substance use. Furthermore, we propose a comprehensive framework for SIP, integrating mental automatism, psychopathological distinctions, and transition mechanisms. This framework aims to refine diagnostic criteria and therapeutic approaches, addressing gaps in clinical practice and research. The study underscores the need for a nuanced understanding of SIP, advocating for a paradigm shift in psychiatric assessment and treatment approaches to better address the complexities of substance-induced mental health disorders.

Seized drug reporting of NPS helps to guide regional toxicological practice: A 17 month review between 2022 and 2023.

Novel psychoactive substances (NPS) are everchanging and plague forensic laboratories who must identify an unending variety of emerging substances and evolve current methodologies to detect these substances. Identifying potential regional NPS targets and timely examining trends in seized drug data could help mitigate the burden laboratories face. Over 17 months, NPS seized drug data were processed and categorized from three laboratories located across the United States to determine any NPS regional similarities and prevalent NPS drug categories: the South Carolina Law Enforcement Division (SLED), the Sedgwick County Regional Forensic Science Center (SCRFSC), and the Orange County Crime Laboratory (OCCL). Seized drug materials, including pills, powders, and plant material, were primarily analyzed for NPS via gas chromatography-mass spectrometry and Fourier transform infrared spectroscopy. From June 2022 to October 2023, 1940 NPS seized drug identifications were reported by these laboratories with 63 different NPS reported. Novel synthetic opioids (NSO) were the most prevalent NPS class across all three laboratories (55%), with fluorofentanyl accounting for 74% of NSO identifications. This is unsurprising given the fentanyl epidemic in the United States. Furthermore, these data highlighted varying regional NPS seized drug trends: eutylone, a synthetic cathinone, was one of the most frequently identified NPS in SLED, SCRFSC observed the most diverse set of synthetic cannabinoids, and OCCL observed an increased prevalence in the designer benzodiazepine, bromazolam. NPS scope recommendations are a valuable resource for forensic laboratories; however, most focus on a national perspective. Timely analysis and reporting of NPS seized drug data may help to develop regional NPS scope recommendations laboratories may employ.

Effects of the Synthetic Cathinone α-Pyrrolidinobutiothiophenone (α-PBT) on Discriminative Stimulus Effects and Intracranial Self-Stimulation Thresholds in Male Rats.

Recently, the abuse of synthetic cathinones is increasing among young people. α-Pyrrolidinobutiothiophenone (α-PBT), a synthetic cathinone, is a designer drug that is freely traded online with no legal restrictions. Moreover, there is currently no scientific basis for legal regulation. Here, we examined the addictive properties of α-PBT using a drug discrimination (DD) task. We also investigated the role of α-PBT in brain stimulation reward (BSR) using an intracranial self-stimulation (ICSS) paradigm in rats. Initially, the rats were trained to discriminate between cocaine and saline. After the discrimination training criteria were met, we determined the dose-effect curves of cocaine and conducted generalization tests with α-PBT and α-pyrrolidinopentiothiophenone (α-PVT) using a cumulative dosing protocol. In a separate set of studies, we examined the dopaminergic mechanisms underlying the function of α-PBT as an interoceptive stimulus (17.8 mg/kg) by intraperitoneally injecting either the dopamine (DA) D1 antagonist SCH23390 (0.06 and 0.12 mg/kg) or the D2 antagonist eticlopride (0.05 and 0.1 mg/kg) 15 min before DD testing. Brain reward function was measured using an ICSS procedure to examine the effects of α-PBT on ICSS threshold under the frequency-rate procedure. Our results showed that α-PBT functioned as a discriminative cue similar to cocaine in rats. More importantly, SCH23390 abolished the effects of α-PBT as an interoceptive stimulus in a dose-dependent manner in rats trained to press a lever to receive cocaine. Similarly, eticlopride dose-dependently attenuated the effect of α-PBT used as a discriminative cue. Additionally, cumulative α-PBT administration dose-dependently lowered ICSS thresholds compared with those in saline-treated rats. Furthermore, α-PBT-induced potentiation of BSR was abolished by pretreatment with both SCH23390 and eticlopride. Taken together, our results suggest that α-PBT can function as a cocaine-like discriminative cue via the activation of D1 and D2 receptors. α-PBT also appears to influence BSR by reducing the brain reward threshold via changes in D1 and D2 receptors. The present study suggests that α-PBT could have addictive properties through DA D1 and D2 receptors and thus poses a threat to humans.

New Psychoactive Substances: Health and Legal Challenges

Drug abuse represents a significant public health problem with a growing tendency. As a way of circumventing the strict national and international control of psychoactive substances by regulatory agencies, there is a market release of new substances with psychoactive activity, called New Psychoactive Substances (NPSs). This group of substances encompasses a diverse range of synthetic compounds designed to mimic the effects of traditional illicit substances. As NPSs show stronger psychoactive effects than classical drugs, they pose unique challenges to public health and regulatory frameworks. Additionally, some substances are considered NPSs in some countries but not in others. Therefore, based on a given legal definition, manufacturers can create an NPS that does not fall under that definition and thus is not prohibited. This review critically explores the multifaceted dimensions of the criminal and legal contexts associated with NPSs. It examines the trends of abuse, the intricate network of criminal and legal aspects surrounding these substances, and the crucial warning signs that indicate their emergence, highlighting the health risks posed by these substances. In conclusion, this manuscript addresses the intricate interplay between the pharmacology, risks, and regulatory responses. These multifaceted challenges associated with NPSs will likely provide valuable insights for future research.

Study of 25R-NBOHs (R = Br, Cl, Et, I): Thermal stability investigation and presumptive identification by SCXRD and PXRD data

New psychoactive substances (NPS) are present in the drug market and can be used as legal substitutes, making enforcement difficult. One of the classes of NPSs is the phenylethylamines, among which 25R-NBOHs (with R being a halogen or an alkyl group) are included. These compounds degrade when subjected to gas chromatography-mass spectrometry (GC–MS), one of the techniques routinely used to identify illicit substances, thereby complicating their identification. This study conducted the identification of NBOH samples using FTIR, XRD, and GC–MS techniques, as well as thermal analysis to obtain information that may assist in the development of new methods for the analysis of these synthetic substances. ATR-FTIR spectra presented characteristic bands of 25R-NBOH compounds (R = Br, Cl, Et, I). The UV–Vis spectra of the 25R-NBOH (R = Br, Cl, I) and 25E-NBOH samples showed a different profile, which may serve as a means to differentiate these compounds. The crystalline structure of the compounds was confirmed by comparing the experimentally obtained diffractograms with the diffraction patterns calculated using data from the Cambridge Crystallographic Data Center (CCDC) database. Thermal analysis indicated that the samples were not pure, confirming the FTIR and XRD analyses, which revealed the presence of impurities and the onset of sample degradation.

The neural basis of resting-state fMRI functional connectivity in fronto-limbic circuits revealed by chemogenetic manipulation

Measures of fMRI resting-state functional connectivity (rs-FC) are an essential tool for basic and clinical investigations of fronto-limbic circuits. Understanding the relationship between rs-FC and the underlying patterns of neural activity in these circuits is therefore vital. Here we introduced inhibitory designer receptors exclusively activated by designer drugs (DREADDs) into the amygdala of two male macaques. We evaluated the causal effect of activating the DREADD receptors on rs-FC and neural activity within circuits connecting amygdala and frontal cortex. Activating the inhibitory DREADD increased rs-FC between amygdala and ventrolateral prefrontal cortex. Neurophysiological recordings revealed that the DREADD-induced increase in fMRI rs-FC was associated with increased local field potential coherency in the alpha band (6.5–14.5 Hz) between amygdala and ventrolateral prefrontal cortex. Thus, our multi-modal approach reveals the specific signature of neuronal activity that underlies rs-FC in fronto-limbic circuits.

A computationally supported designer benzodiazepine strategy for public toxicology laboratories.

Public laboratories must balance innovative and existing methods to keep up with designer drug trends. This article presents a strategy for handling designer benzodiazepines (DBZDs) in casework from screening to interpretation. The cross-reactivity of 22 DBZDs and metabolites was tested against the Immunalysis™ Benzodiazepine Direct Enzyme-Linked Immunosorbent Assay kit. The kit had high intra-analyte precision (coefficients of variation < 15%). Inter-analyte performance varied, triggering confirmation testing at concentrations ranging from 35 to 460 μg/L. The CCRFSL implemented a 40-analyte benzodiazepine and Z-drug confirmation method in 2019. Ten additional analytes were later validated for qualitative reporting, and the limits of detection (LODs) for 13 analytes were lowered by 60%. The method of standard addition was also optimized for as-needed quantitation. Equal and 1/x weighting factors correlated well with target concentrations (coefficients of determination (r2) > 0.98), but 1/x weighting provided the most consistently accurate concentrations. Six computational models were developed to predict DBZD binding affinity to the γ-aminobutyric acid-A receptor to assist in case interpretation (r2 > 0.7 for cross-validation and test set prediction). These models were used to predict the binding affinity of analytes in the confirmation method. Other public laboratories can use this same practical strategy to adapt to any designer drug class (e.g., benzodiazepines, opioids, cannabinoids, and stimulants).

Toxicity of the New Psychoactive Substance (NPS) Clephedrone (4-Chloromethcathinone, 4-CMC): Prediction of Toxicity Using In Silico Methods for Clinical and Forensic Purposes.

This study reports the first application of in silico methods to assess the toxicity of 4-chloromethcathinone (4-CMC), a novel psychoactive substance (NPS). Employing advanced toxicology in silico tools, it was possible to predict crucial aspects of the toxicological profile of 4-CMC, including acute toxicity (LD50), genotoxicity, cardiotoxicity, and its potential for endocrine disruption. The obtained results indicate significant acute toxicity with species-specific variability, moderate genotoxic potential suggesting the risk of DNA damage, and a notable cardiotoxicity risk associated with hERG channel inhibition. Endocrine disruption assessment revealed a low probability of 4-CMC interacting with estrogen receptor alpha (ER-α), suggesting minimal estrogenic activity. These insights, derived from in silico studies, are critical in advancing the understanding of 4-CMC properties in forensic and clinical toxicology. These initial toxicological findings provide a foundation for future research and aid in the formulation of risk assessment and management strategies in the context of the use and abuse of NPSs.

Dorsal raphe to basolateral amygdala corticotropin-releasing factor circuit regulates cocaine-memory reconsolidation.

Environmental stimuli elicit drug craving and relapse in cocaine users by triggering the retrieval of strong cocaine-related contextual memories. Retrieval can also destabilize drug memories, requiring reconsolidation, a protein synthesis-dependent storage process, to maintain memory strength. Corticotropin-releasing factor (CRF) signaling in the basolateral amygdala (BLA) is necessary for cocaine-memory reconsolidation. We have hypothesized that a critical source of CRF in the BLA is the dorsal raphe nucleus (DR) based on its neurochemistry, anatomical connectivity, and requisite involvement in cocaine-memory reconsolidation. To test this hypothesis, male and female Sprague-Dawley rats received adeno-associated viruses to express Gi-coupled designer receptors exclusively activated by designer drugs (DREADDs) selectively in CRF neurons of the DR and injection cannulae directed at the BLA. The rats were trained to self-administer cocaine in a distinct environmental context then received extinction training in a different context. Next, they were briefly re-exposed to the cocaine-predictive context to destabilize (reactivate) cocaine memories. Intra-BLA infusions of the DREADD agonist deschloroclozapine (DCZ; 0.1 mM, 0.5 µL/hemisphere) immediately after memory reactivation attenuated cocaine-memory strength, relative to vehicle infusion. This was indicated by a selective, DCZ-induced and memory reactivation-dependent decrease in drug-seeking behavior in the cocaine-predictive context in DREADD-expressing males and females at test compared to respective controls. Notably, BLA-projecting DR CRF neurons that exhibited increased c-Fos expression during memory reconsolidation co-expressed the glutamatergic neuronal marker, vesicular glutamate transporter 3. Together, these findings suggest that the DRCRF → BLA circuit is engaged to maintain cocaine-memory strength after memory destabilization, and this phenomenon may be mediated by DR CRF and/or glutamate release in the BLA.

N-Ethylhexedrone: A very long and bad trip! A case series.

N-ethylhexedrone (NEH) is a new cathinone derivative with, currently, low toxicokinetic and toxicodynamic knowledge. We present three documented clinical cases of NEH intoxication with plasma and urine concentrations. A thorough search for metabolites was performed. The three patients were admitted to the emergency department, and two out of the three were hospitalized for an extended period. While recovering from the drug effects, 12-24 h after nasal intake of New Psychoactive Substance (NPS), the patients described the following disorders: anxiety, feelings of persecution, asthenia, anhedonia, abulia, psychomotor slowing and loss of consciousness. NEH was identified in all samples by liquid chromatography-high resolution mass spectrometry (LC-HRMS), and quantified by liquid chromatography coupled to tandem mass spectrometry (LC-MS-MS). Quantitative analysis showed decreasing concentrations over time: for Case 1, from 97.2 (Day 1, D1) to 0.7 (Day 7, D7) µg/L for plasma, and from 724 (D1) to 0.5 (D7) µg/L for urine. NEH concentration of 7.9 µg/L was found in the plasma collected at admission for Case 2. For Case 3, concentrations ranging from 49 (D1) to 1.8 (D7) µg/L in plasma, and from 327.3 (Day 6, D6) to 116.8 (D7) µg/L in urine were found. NEH was no longer detected in the urine sample at Day 10. Elimination half-life was estimated at 19, and 28 hours in Patients 1 and 3, respectively. Four metabolites were identified in blood and urine: reduced NEH, dealkyl-NEH, reduced dealkyl-NEH and hydroxy-NEH. The cases presented highlight the long detectable lifetime of NEH. Characterization of the metabolites will allow better identification of the consumption of this drug. Serious adverse events can be observed after NEH consumption, as two out of the three patients required intubation and ventilation. A syndrome of inappropriate antidiuretic hormone secretion (SIADH) was also diagnosed. Two out of the three cases are notable because of the number of samples collected and because NEH was the only drug of abuse detected.

Structure-Activity Relationships, Deuteration, and Fluorination of Synthetic Cannabinoid Receptor Agonists Related to AKB48, 5F-AKB-48, and AFUBIATA.

Synthetic cannabinoid receptor agonists (SCRAs) are a growing class of new psychoactive substances (NPS) commonly derived from an N-alkylated indole, indazole, or 7-azaindole scaffold. Diversification of this core (at the 3-position) with amide-linked pendant amino acid groups and modular N-alkylation (of the indole/indazole/7-azaindole core) ensures that novel SCRAs continue to enter the illicit drug market rapidly. In response to the large number of SCRAs that have been detected, pharmacological evaluation of this NPS class has become increasingly common. Adamantane-derived SCRAs have consistently appeared throughout the market since 2011, and as such, a systematic set of these derivatives was synthesized and pharmacologically evaluated. Deuterated and fluorinated adamantane derivatives were prepared to evaluate typical hydrogen bioisosteres, as well as evaluation of the newly detected AFUBIATA.

Disrupting dorsal hippocampus impairs category learning in rats

Categorization requires a balance of mechanisms that can generalize across common features and discriminate against specific details. A growing literature suggests that the hippocampus may accomplish these mechanisms by using fundamental mechanisms like pattern separation, pattern completion, and memory integration. Here, we assessed the role of the rodent dorsal hippocampus (HPC) in category learning by combining inhibitory DREADDs (Designer Receptors Exclusively Activated by Designer Drugs) and simulations using a neural network model. Using touchscreens, we trained rats to categorize distributions of visual stimuli containing black and white gratings that varied along two continuous dimensions. Inactivating the dorsal HPC impaired category learning and generalization, suggesting that the rodent HPC plays an important role during categorization. Hippocampal inactivation had no effect on a control discrimination task that used identical trial procedures as the categorization tasks, suggesting that the impairments were specific to categorization. Model simulations were conducted with variants of a neural network to assess the impact of selective deficits on category learning. The hippocampal inactivation groups were best explained by a model that injected random noise into the computation that compared the similarity between category stimuli and existing memory representations. This model is akin to a deficit in mechanisms of pattern completion, which retrieves similar memory representations using partial information.

Evaluation of Density Functional Theory-Generated Data for Infrared Spectroscopy of Novel Psychoactive Substances Using Unsupervised Learning

Novel psychoactive substances (NPSs) are compounds plotted to modify the chemical structures of prohibited substances, offering alternatives for consumption and evading legislation. The prompt emergence of these substances presents challenges in health concerns and forensic assessment because of the lack of analytical standards. A viable alternative for establishing these standards involves leveraging in silico methods to acquire spectroscopic data. This study assesses the efficacy of utilizing infrared spectroscopy (IRS) data derived from density functional theory (DFT) for analyzing NPSs. Various functionals were employed to generate infrared spectra for five distinct NPS categories including the following: amphetamines, benzodiazepines, synthetic cannabinoids, cathinones, and fentanyls. PRISMA software was conceived to rationalize data management. Unsupervised learning techniques, including Hierarchical Cluster Analysis (HCA), Principal Component Analysis (PCA), and t-distributed stochastic neighbor embedding (t-SNE), were utilized to refine the assessment process. Our findings reveal no significant disparities among the different functionals used to generate infrared spectra data. Additionally, the application of unsupervised learning demonstrated adequate segregation of NPSs within their respective groups. In conclusion, integrating theoretical data and dimension reduction techniques proves to be a powerful strategy for evaluating the spectroscopic characteristics of NPSs. This underscores the potential of this combined methodology as a diagnostic tool for distinguishing IR spectra across various NPS groups, facilitating the evaluation of newly unknown compounds.

Identification of Gα12-vs-Gα13-coupling determinants and development of a Gα12/13-coupled designer GPCR

G-protein-coupled receptors (GPCRs) transduce diverse signals into the cell by coupling to one or several Gα subtypes. Of the 16 Gα subtypes in human cells, Gα12 and Gα13 belong to the G12 subfamily and are reported to be functionally different. Notably, certain GPCRs display selective coupling to either Gα12 or Gα13, highlighting their significance in various cellular contexts. However, the structural basis underlying this selectivity remains unclear. Here, using a Gα12-coupled designer receptor exclusively activated by designer drugs (DREADD; G12D) as a model system, we identified residues in the α5 helix and the receptor that collaboratively determine Gα12-vs-Gα13 selectivity. Residue-swapping experiments showed that G12D distinguishes differences between Gα12 and Gα13 in the positions G.H5.09 and G.H5.23 in the α5 helix. Molecular dynamics simulations observed that I378G.H5.23 in Gα12 interacts with N1032.39, S1693.53 and Y17634.53 in G12D, while H364G.H5.09 in Gα12 interact with Q2645.71 in G12D. Screening of mutations at these positions in G12D identified G12D mutants that enhanced coupling with Gα12 and to an even greater extent with Gα13. Combined mutations, most notably the dual Y17634.53H and Q2645.71R mutant, further enhanced Gα12/13 coupling, thereby serving as a potential Gα12/13-DREADD. Such novel Gα12/13-DREADD may be useful in future efforts to develop drugs that target Gα12/13 signaling as well as to identify their therapeutic indications.

How Does the Inner Retinal Network Shape the Ganglion Cells Receptive Field? A Computational Study.

We consider a model of basic inner retinal connectivity where bipolar and amacrine cells interconnect and both cell types project onto ganglion cells, modulating their response output to the brain visual areas. We derive an analytical formula for the spatiotemporal response of retinal ganglion cells to stimuli, taking into account the effects of amacrine cells inhibition. This analysis reveals two important functional parameters of the network: (1) the intensity of the interactions between bipolar and amacrine cells and (2) the characteristic timescale of these responses. Both parameters have a profound combined impact on the spatiotemporal features of retinal ganglion cells' responses to light. The validity of the model is confirmed by faithfully reproducing pharmacogenetic experimental results obtained by stimulating excitatory DREADDs (Designer Receptors Exclusively Activated by Designer Drugs) expressed on ganglion cells and amacrine cells' subclasses, thereby modifying the inner retinal network activity to visual stimuli in a complex, entangled manner. Our mathematical model allows us to explore and decipher these complex effects in a manner that would not be feasible experimentally and provides novel insights in retinal dynamics.

In Vivo and In Vitro Metabolic Fate and Urinary Detectability of Five Deschloroketamine Derivatives Studied by Means of Hyphenated Mass Spectrometry.

Ketamine derivatives such as deschloroketamine and deschloro-N-ethyl-ketamine show dissociative and psychoactive properties and their abuse as new psychoactive substances (NPSs) has been reported. Though some information is available on the biotransformation of dissociative NPSs, data on deschloro-N-cyclopropyl-ketamine deschloro-N-isopropyl-ketamine and deschloro-N-propyl-ketamine concerning their biotransformation and, thus, urinary detectability are not available. The aims of the presented work were to study the in vivo phase I and II metabolism; in vitro phase I metabolism, using pooled human liver microsomes (pHLMs); and detectability, within a standard urine screening approach (SUSA), of five deschloroketamine derivatives. Metabolism studies were conducted by collecting urine samples from male Wistar rats over a period of 24 h after their administration at 2 mg/kg body weight. The samples were analyzed using liquid chromatography high-resolution tandem mass spectrometry (LC-HRMS/MS) and gas chromatography-mass spectrometry (GC-MS). The compounds were mainly metabolized by N-dealkylation, hydroxylation, multiple oxidations, and combinations of these metabolic reactions, as well as glucuronidation and N-acetylation. In total, 29 phase I and 10 phase II metabolites were detected. For the LC-HRMS/MS SUSA, compound-specific metabolites were identified, and suitable screening targets could be recommended and confirmed in pHLMs for all derivatives except for deschloro-N-cyclopropyl-ketamine. Using the GC-MS-based SUSA approach, only non-specific acetylated N-dealkylation metabolites could be detected.

Screening of Astec® CHIRALDEX™ G-PN and LIPODEX™ D gas chromatography columns for enantioseparation of amphetamine derivatives.

Among different substance classes, New Psychoactive Substances (NPS) comprise chiral amphetamines for stimulant and empathic effects. There is little knowledge in terms of clinical studies about possibly different effects of the two enantiomers of novel amphetamine derivatives. For this reason, there is a big demand for enantioseparation method development of this new substance class. Regarding gas chromatography, cyclodextrins proved to be effective for enantioseparation of NPS. In our attempt, an Astec® Chiraldex™ G-PN column containing 2,6-di-O-pentyl-3-propionyl-γ-cyclodextrin and a Lipodex™ D column containing heptakis-(2,6-di-O-pentyl-O-acetyl)-β-cyclodextrin as chiral selector served as stationary phases in a Shimadzu GCMS-QP2010 SE system. Because of the special coating, maximum temperature is limited to 200 °C isothermal or 220 °C in programmed mode. To ensure detection, trifluoroacetic anhydride (TFAA) was used to increase sample volatility.1 As a result, 35 amphetamines were tested as their TFAA-derivatives. A screening method with a temperature gradient from 140 °C to 200 °C at a heating ramp of 1°C per minute and final time of 5min, showed baseline separation for seven and partial separations for 16 trifluoro acetylated amphetamines using the Chiraldex™ G-PN column. Six baseline and nine partial separations were observed with the Lipodex™ D column, respectively.

Targeting Melanocortin 4 Receptor to Treat Sleep-Disordered Breathing

There is no effective pharmacotherapy for sleep-disordered breathing (SDB). A melanocortin receptor 4 (MCR4) agonist, setmelanotide (SET), is used to treat genetic obesity caused by abnormal melanocortin and leptin signaling. We hypothesized that SET can treat SDB in diet-induced obese mice. We performed a proof-of-concept randomized crossover trial of a single dose of SET vs vehicle and a two-week daily SET vs vehicle trial in obese mice. We also examined co-localization of Mcr4 mRNAs with a marker of CO2 sensing neurons PHOX2b in the brainstem and performed chemogenetic studies expressing Cre-dependent designer receptors exclusively activated by designer drugs (DREADD) in Mcr4-Cre mice. SET increased minute ventilation across sleep/wake states, greatly enhanced the hypercapnic ventilatory response (HCVR), and abolished apneas during sleep. PHOX2b+ neurons in the nucleus of the solitary tract (NTS) and the parafacial region expressed Mc4r. Chemogenetic stimulation of the MC4R+ neurons in the parafacial region, but not in the NTS, augmented the HCVR without any changes in metabolism. Parafacial MC4R neurons projected to the respiratory pre-motor neurons expressing cholera toxin B after C3-C4 spinal cord injections. In conclusion, MC4R agonists enhance the HCVR and treat SDB by acting on the parafacial MC4R+ neurons. NIH (R01 HL128970, R01 HL133100, and R01 HL138932), AHA (#915167). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Inhibition of somatostatinergic interneurons of the lateral parafacial region reduces the ventilatory response to hypercapnia of mice

The lateral parafacial region (pFL), which contains excitatory and inhibitory neurons and interneurons, generates active expiration in response to hypercapnia. Inhibition of inhibitory somatostatinergic (Sst)-positive neurons in parafacial region increases pulmonary ventilation, while hypercapnia inhibits some of them intrinsically. However, the role of pFL Sst-positive interneurons (with local projections) in regulating pulmonary ventilation during hypercapnia is unclear. We hypothesized that the pFL Sst-positive interneurons control the pulmonary ventilation of mice under normocapnia and hypercapnia, as part of a local neural network. Herein, we evaluated the impact of pFL Sst-positive interneurons inhibition on pulmonary ventilation of non-anesthetized mice during normocapnia and hypercapnia. We used genetically modified mice, expressing the cre recombinase in Sst-positive neurons, and injected unilaterally a cre-dependent adeno-associated (AAV) retrograde virus in the pFL region to express the FLP recombinase, followed by a cre- and FLP-dependent AAV to express the inhibitory Designer Receptors Exclusively Activated by Designer Drugs (hM4D-Gi) in the pFL Sst-positive interneurons. We used whole-body plethysmography to measure the respiratory frequency (fR), tidal volume (VT), pulmonary ventilation (VE), as well as the duration of the respiratory cycle, inspiration (DI) and expiration (DE) at normocapnia and hypercapnia (7% CO2) after activation of hM4D-Gi in the pFL Sst-positive interneurons using JHU 37160 (JHU; 0.1 mg/kg; i.p.). Data are expressed as mean ± standard deviation. By analyzing the expression of mCherry fluorescent protein, we found a significant number of Sst-positive interneurons (73.43 ± 10.52) in the pFL region (n=14). JHU administration did not affect the ventilatory parameters of Sst(cre/cre) (n= 9) and Sst(cre/cre + hM4D-Gi) (n= 14) groups at normocapnia [(Δ fR: 6.5 ± 33.56 vs. -14.86 ± 49.03 cpm; p= 0.42) (Δ VT: 0.87 ± 1.15 vs. -0.97 ± 2.58 μL·g−1; p= 0.19) (Δ VE: 235.8 ± 388.8 vs. -315.9 ± 725.1 μL·g−1·min−1; p= 0.16) (Δ duration of the respiratory cycle: -13 ± 45.11 vs. 12.34 ± 53.07 ms; p= 0.39) (Δ DI: 10.36 ± 11.23 vs. 14.7 ± 16.38 ms; p= 0.62) (Δ DE: -23.36 ± 42.42 vs. -2.41± 53.58 ms; p= 0.48)]. On the other hand, JHU administration attenuated the hypercapnia-induced responses of all ventilatory parameters, such as fR (Δ: 44.57 ± 56.66 vs. 126± 21.28 cpm; p= 0.0002), VT (Δ: 1.32 ± 3.35 vs. 4.16 ± 2.56 μL·g−1; p= 0.04), VE (Δ: 948.7 ± 1407.1 vs. 2183 ± 909.4 μL·g−1·min−1; p= 0.03), duration of the respiratory cycle (Δ: -44.57 ± 46.31 vs. -122.1 ± 30.7 ms; p= 0.0002), DI (Δ: 6.79 ± 16.36 vs. -13.03 ± 10.19 ms; p= 0.003) and DE (Δ: -51.37 ± 37.50 vs. -109.1 ± 32.78 ms; p= 0.001) in the Sst(cre/cre + hM4D-Gi) compared with the Sst(cre/cre) group. Thus, we conclude that the pFL Sst-positive interneurons control the ventilatory parameters of mice during hypercapnia, but not during normocapnia. FAPESP, CNPq and CAPES. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.