Structural and functional perspectives on interactions between synthetic cathinones and monoamine transporters.

Synthetic cathinone derivatives comprise a family of psychoactive compounds structurally related to amphetamine. Over the last decade, clandestine chemists have synthesized a consistent stream of innovative cathinone derivatives to outpace governmental regulatory restrictions. Many of these unregulated substances are produced and distributed as designer drugs. Two of the principal chemical scaffolds exploited to expand the synthetic cathinone family are methcathinone and α-pyrrolidinopentiophenone (or α-pyrrolidinovalerophenone, α-PVP). These compounds' main physiological targets are monoamine transporters, where they promote addiction by potentiating dopaminergic neurotransmission. This chapter describes techniques used to study the pharmacodynamic properties of cathinones at monoamine transporters in vitro. Biochemical techniques described include uptake inhibition and release assays in rat brain synaptosomes and in mammalian expression systems. Electrophysiological techniques include current measurements using the voltage clamp technique. We describe a Ca2+ mobilization assay wherein voltage-gated Ca2+ channels function as reporters to study the action of synthetic cathinones at monoamine transporters. We discuss results from systematic structure-activity relationship studies on simple and complex cathinones at monoamine transporters with an emphasis on identifying structural moieties that modulate potency and selectivity at these transporters. Moreover, different profiles of selectivity at monoamine transporters directly predict compounds associated with behavioral and subjective effects within animals and humans. In conclusion, clarification of the structural aspects of compounds which modulate potency and selectivity at monoamine transporters is critical to identify and predict potential addictive drugs. This knowledge may allow prompt allocation of resources toward drugs that represent the greatest threats after drugs are identified by forensic laboratories.

Concussive Head Trauma Deranges Axon Initial Segment Function in Axotomized and Intact Layer 5 Pyramidal Neurons.

The axon initial segment (AIS) is a critical locus of control of action potential (AP) generation and neuronal information synthesis. Concussive traumatic brain injury gives rise to diffuse axotomy, and the majority of neocortical axonal injury arises at the AIS. Consequently, concussive traumatic brain injury might profoundly disrupt the functional specialization of this region. To investigate this hypothesis, one and two days after mild central fluid percussion injury in Thy1-YFP-H mice, we recorded high-resolution APs from axotomized and adjacent intact layer 5 pyramidal neurons and applied a second derivative (2o) analysis to measure the AIS- and soma-regional contributions to the AP upstroke. All layer 5 pyramidal neurons recorded from sham animals manifested two stark 2o peaks separated by a negative intervening slope. In contrast, within injured mice, we discovered a subset of axotomized layer 5 pyramidal neurons in which the AIS-regional 2o peak was abolished, a functional perturbation associated with diminished excitability, axonal sprouting and distention of the AIS as assessed by staining for ankyrin-G. Our analysis revealed an additional subpopulation of both axotomized and intact layer 5 pyramidal neurons that manifested a melding together of the AIS- and soma-regional 2o peaks, suggesting a more subtle aberration of sodium channel function and/or translocation of the AIS initiation zone closer to the soma. When these experiments were repeated in animals in which cyclophilin-D was knocked out, these effects were ameliorated, suggesting that trauma-induced AIS functional perturbation is associated with mitochondrial calcium dysregulation.

Somatostatin interneurons exhibit enhanced functional output and resilience to axotomy after mild traumatic brain injury.

Mild traumatic brain injury (mTBI) gives rise to a remarkable breadth of pathobiological consequences, principal among which are traumatic axonal injury and perturbation of the functional integrity of neuronal networks that may arise secondary to the elimination of the presynaptic contribution of axotomized neurons. Because there exists a vast diversity of neocortical neuron subtypes, it is imperative to elucidate the relative vulnerability to axotomy among different subtypes. Toward this end, we exploited SOM-IRES-Cre mice to investigate the consequences of the central fluid percussion model of mTBI on the microanatomical integrity and the functional efficacy of the somatostatin (SOM) interneuron population, one of the principal subtypes of neocortical interneuron. We found that the SOM population is resilient to axotomy, representing only 10% of the global burden of inhibitory interneuron axotomy, a result congruous with past work demonstrating that parvalbumin (PV) interneurons bear most of the burden of interneuron axotomy. However, the intact structure of SOM interneurons after injury did not translate to normal cellular function. One day after mTBI, the SOM population is more intrinsically excitable and demonstrates enhanced synaptic efficacy upon post-synaptic layer 5 pyramidal neurons as measured by optogenetics, yet the global evoked inhibitory tone within layer 5 is stable. Simultaneously, there exists a significant increase in the frequency of miniature inhibitory post-synaptic currents within layer 5 pyramidal neurons. These results are consistent with a scheme in which 1 day after mTBI, SOM interneurons are stimulated to compensate for the release from inhibition of layer 5 pyramidal neurons secondary to the disproportionate axotomy of PV interneurons. The enhancement of SOM interneuron intrinsic excitability and synaptic efficacy may represent the initial phase of a dynamic process of attempted autoregulation of neocortical network homeostasis secondary to mTBI.

Broad, Multi-Year Sampling Effort Highlights Complex Dynamics of the Tick-Borne Pathogen Ehrlichia chaffeensis (Rickettsiales: Anaplasmatacae).

Ehrlichia chaffeensis (Rickettsiales: Anaplasmatacae), an understudied bacterial pathogen emerging in the eastern United States, is increasing throughout the range of its vector, the lone star tick [Amblyomma americanum, L. (Acari: Ixodidae)]. To mitigate human disease risk, we must understand what factors drive E. chaffeensis prevalence. Here, we report patterns of E. chaffeensis prevalence in southeastern Virginia across 4 yr and ask how seasonal weather patterns affect variation in rates of E. chaffeensis occurrence. We collected A. americanum nymphs at 130 plots across southeastern Virginia in 2012, 2013, 2015, and 2016, and used polymerase chain reaction and gel electrophoresis to test for the presence of E. chaffeensis DNA. Prevalence estimates varied among years, ranging from 0.9% to 3.7%, and persistence of E. chaffeensis occurrence varied across space, with some sites never testing positive, and one site testing positive every year. Using generalized linear mixed-effects models, we related E. chaffeensis occurrence to temperature, humidity, vapor-pressure deficit, and precipitation during seasons up to 21 mo prior to sampling. Surprisingly, all support was lent to a positive effect of temperature during the previous fall and winter (i.e., prior to the nymphs' hatching), which we hypothesize to influence reservoir host population dynamics through changes to mortality or natality. Although further work is necessary to truly elucidate the mechanisms at play, our study shows E. chaffeensis distribution to be very dynamic across multiple dimensions, demanding broad concerted monitoring efforts that can consider both space and time.