Angela L. Purcell

Rarely a new research area has gotten such an overwhelming amount of attention as have microRNAs. Although several basic questions regarding their biological principles still remain to be answered, many specific characteristics of microRNAs in combination with compelling therapeutic efficacy data and a clear involvement in human disease have triggered the biotechnology community to start exploring the possibilities of viewing microRNAs as therapeutic entities. This review serves to provide some general insight into some of the current microRNAs targets, how one goes from the initial bench discovery to actually developing a therapeutically useful modality, and will briefly summarize the current patent landscape and the companies that have started to explore microRNAs as the next drug target.

Using the confocal microscope together with markers for the cellular components of glomeruli, we examined the spatiotemporal cellular interactions that occur between the axons of olfactory receptor cells, their dendritic targets, and glial cells during the critical period of glomerular formation. We have employed markers of immature and mature olfactory receptor cell axons, mitral/tufted cell dendrites, and glial cells as well as a synapse-associated protein for double- and triple-label immunocytochemistry. Axons of olfactory receptor cells grew into a dense dendritic zone of the olfactory bulb (comprising the dendrites of both mitral and tufted cells) between E17 and E18. At E19, these axons coalesced into protoglomeruli, which continued to develop until birth, when the basic anatomical structure of adult glomeruli emerged. Neither mitral/tufted cell dendrites nor olfactory bulb astrocytes became specifically associated with these protoglomeruli until E21. Ensheathing cells remained restricted to the outer nerve fiber layer and did not appear to contribute to glomerular formation. Finally, the synaptophysin staining has shown that synaptic constituents are expressed as early as E17, prior to the appearance of mature olfactory receptor cell axons. Based on these data, we have established a time line detailing the temporal and spatial interactions that occur between cell types during late embryonic rat olfactory bulb development. We conclude that the initial event in the formation of glomeruli is the penetration of the mitral/tufted cell dendritic zone by olfactory receptor cell axons. The coalescence of dendritic and glial processes into glomerular structures appears secondary to the arrival of the olfactory receptor cell axons.

Nerve-evoked contractions were studied in vitro in phrenic nerve-hemidiaphragm preparations from strain 129X1 acetylcholinesterase knockout (AChE-/-) mice and their wild-type littermates (AChE+/+). The AChE-/- mice fail to express AChE but have normal levels of butyrylcholinesterase (BChE) and can survive into adulthood. Twitch tensions elicited in diaphragms of AChE-/- mice by single supramaximal stimuli had larger amplitudes and slower rise and decay times than did those in wild-type animals. In AChE-/- preparations, repetitive stimulation at frequencies of 20 and 50 Hz and at 200 and 400 Hz produced decremental muscle tensions; however, stimulation at 70 and 100 Hz resulted in little or no loss of tension during trains. Muscles from AChE+/+ mice maintained tension at all frequencies examined but exhibited tetanic fade after exposure to the selective AChE inhibitor 1,5-bis(4-allyldimethyl-ammoniumphenyl)pentane-3-one (BW 284C51). The ability of diaphragm muscles from AChE-/- mice to maintain tension at 70 and 100 Hz suggests a partial compensation for impairment of acetylcholine (ACh) hydrolysis. Three mechanisms--including a reliance on BChE activity for termination of ACh action, downregulation of nicotinic acetylcholine receptors (nAChRs), and morphological remodeling of the endplate region--were identified. Studies of neuromuscular transmission in this model system provide an excellent opportunity to evaluate the role of AChE without complications arising from use of inhibitors.