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Applying voltage-imaging techniques to visualise the effect of lysergic acid diethylamide (LSD) on cortical layer V pyramidal cells and its adaptation during tolerance development

Periodic Reporting for period 1 - LSD-CortLVPC (Applying voltage-imaging techniques to visualise the effect of lysergic acid diethylamide (LSD) on cortical layer V pyramidal cells and its adaptation during tolerance development)

Reporting period: 2016-06-01 to 2018-05-31

Serotonergic hallucinogens, such as lysergic acid diethylamide (LSD), are psychoactive drugs which induce profound alterations of consciousness. They are popular drugs of abuse, but most recently also re-gained attention as possible promotors of psychotherapy. The controversy surrounding these drugs remains unresolved by modern science and politics, and demands for a strong scientific basis able to reveal how these drugs work and how their working potentially translates into benefit and/or detriment to human health and society.
Serotonergic hallucinogens interact with a variety of different monoamine receptors (proteins that govern the communication patterns across neurons), yet their psychoactive effects are thought to be primarily mediated by serotonin (5-HT) 2A receptors. 5-HT2A receptors densely populate certain neurons within the cortex of the brain (particularly influencing so called pyramidal cells), but they can also be found widely across the vascular system. How neuronal and vascular 5-HT2A receptors orchestrate to give rise to the brain dynamics characteristic for these drugs, however, is still largely unknown. That most serotonergic hallucinogens (including LSD) tend to be highly promiscuous in their receptor binding preferences, adds another layer of complexity and renders any final appraisal of the mechanistic implications of 5-HT2A receptors difficult. Here, taking advantage of a recent development in drug design, we use the highly selective 5-HT2(A) agonist 25CN-NBOH in mice. We describe the drug as to the essentials of its behavioural and autonomic properties; apply pulse oximetry to measure drug induced blood flow changes in brain-imminent arteries; and follow up on peripyramidal haemodynamics and midlayer pyramidal cell activity by means of cutting-edge techniques of optogenetic brain imaging. The overall aim is to provide insights into the (neuro-)physiology of 5-HT2A activity, delineating a possible mechanism of action of serotonergic hallucinogens.
With the most important results of our project we find that 25CN-NBOH induces head twitches in mice, a benign behaviour characteristic for hallucinogen action in rodents, which is sensitive to pharmacological blockade of 5-HT2A receptors, and which shows rapid (hallucinogen-typical) development of tolerance. 25CN-NBOH, as indicated by neck-arterial pulse oximetry and body core temperature measurements, modulates breathing and thermoregulation, and increases heart rate as well as pulse distention (an index of blood flow) in brain-imminent neck arteries. The effect of 25CN-NBOH on neck (most likely carotid) arteries is particularly pronounced when the body-warming effect of the drug is facilitated by environmental heat. Genetic targeting of the refined voltage-sensitive fluorescent protein butterfly 1.2. to midlayer pyramidal cells of the cortex reveals that 25CN-NBOH too interferes with the peripyramidal haemodynamics of the brain. In particular, we find that the drug leads to a dispersion of pulse-driven blood flow related signal oscillations (BfSOs) and reduces their frequency and strength. For investigation of a possible role of motor-cortical pyramidal cells for 25CN-NBOH induced head twitches, we furthermore used VSFP-emission synchronised body-close-up video recordings to monitor the animals’ behaviour along with their neuronal activity. We demonstrate that 25CN-NBOH induced head twitches have a strong gamma frequency component. First (preliminary) analysis of the associated pyramidal VSFP emissions seems to indicate that there might also be a band-specific imbedding of the given behaviour in the cortex. Additional analysis and complementary research addressing the cortical components of 5-HT2A related motor output appear warranted. For further information, feel invited to have a look at the first-delivery open-access paper of our project (https://doi.org/10.3389/fphar.2018.00017) and to follow up on our publication updates on the project related webpage (www.psyborn.com/msca-blog).
Research on the mechanism of action of serotonergic hallucinogens is weakened by the unselectivity of most of the drugs of this class and a lack of methods allowing to discriminate neuronal vs. haemodynamic signals associated with cortical 5-HT2A agonism. We are amongst the first to introduce 25CN-NBOH as a new key player of hallucinogen research and provide the very essentials of the drug’s behavioural, autonomic, and cardiovascular profile. Understanding hallucinogens not only as to neuronal activity but also as to neuronal-activity-sustaining haemodynamics is a major challenge of contemporary neuroscience. As human research on these drugs heavily is reliant on fMRI and PET methodology, disentangling peripheral, brain-imminent and brain-intrinsic sites of vascular hallucinogen action seems pivotal. Showing that 25CN-NBOH affects heart rate, carotid as well peripyramidal blood flow in the cortex might help to provide a more global picture on hallucinogen physiology and in the context of thermoregulation moreover open up a mechanistic framework. As there is an increasing scientific movement towards reintegration of certain serotonergic hallucinogens into psychotherapy on the one hand, and the potential for abuse and harm in unsupervised contexts on the other, learning about these drugs appears to have become more important than ever.
Effect of 25CN-NBOH on neck-arterial and peripyramidal haemodynamics