Graded and discontinuous EphA–ephrinB expression patterns in the developing auditory brainstem

Abstract

Eph-ephrin interactions guide topographic mapping and pattern formation in a variety of systems. In contrast to other sensory pathways, their precise role in the assembly of central auditory circuits remains poorly understood. The auditory midbrain, or inferior colliculus (IC) is an intriguing structure for exploring guidance of patterned projections as adjacent subdivisions exhibit distinct organizational features. The central nucleus of the IC (CNIC) and deep aspects of its neighboring lateral cortex (LCIC, Layer 3) are tonotopically-organized and receive layered inputs from primarily downstream auditory sources. While less is known about more superficial aspects of the LCIC, its inputs are multimodal, lack a clear tonotopic order, and appear discontinuous, terminating in modular, patch/matrix-like distributions. Here we utilize X-Gal staining approaches in lacZ mutant mice (ephrin-B2, -B3, and EphA4) to reveal EphA-ephrinB expression patterns in the nascent IC during the period of projection shaping that precedes hearing onset. We also report early postnatal protein expression in the cochlear nuclei, the superior olivary complex, the nuclei of the lateral lemniscus, and relevant midline structures. Continuous ephrin-B2 and EphA4 expression gradients exist along frequency axes of the CNIC and LCIC Layer 3. In contrast, more superficial LCIC localization is not graded, but confined to a series of discrete ephrin-B2 and EphA4-positive Layer 2 modules. While heavily expressed in the midline, much of the auditory brainstem is devoid of ephrin-B3, including the CNIC, LCIC Layer 2 modular fields, the dorsal nucleus of the lateral lemniscus (DNLL), as well as much of the superior olivary complex and cochlear nuclei. Ephrin-B3 LCIC expression appears complementary to that of ephrin-B2 and EphA4, with protein most concentrated in presumptive extramodular zones. Described tonotopic gradients and seemingly complementary modular/extramodular patterns suggest Eph-ephrin guidance in establishing juxtaposed continuous and discrete neural maps in the developing IC prior to experience.

Modular‐extramodular organization in developing multisensory shell regions of the mouse inferior colliculus

“…Very little is known concerning the developmental timing and shaping of multimodal axonal patterns in the LCIC. Since compartmental Eph-ephrin LCIC expression patterns correlate temporally with early CNIC guidance gradients (Gabriele et al, 2011;Wallace et al, 2013Wallace et al, , 2016, we anticipate a similar time course for LCIC modularextramodular circuit assembly as has been previously documented for developing CNIC afferent layers (Fathke & Gabriele, 2009; Gabriele, -Bechtold, & Henkel, 2000a,b;Gabriele, Shahmoradian, French, Henkel, & McHaffie, 2007;Gabriele et al, 2011;Wallace et al, 2013).…”

“…Both the modular patterning revealed through AChE, NADPH-d, GAD, and CO staining, as well as the highly specific CR extramodular labeling, is reminiscent of previous reports from our laboratory in neonatal mouse and rat highlighting early LCIC guidance expression patterns (Cramer & Gabriele, 2014;Gabriele et al, 2011;Wallace, Kavianpour, & Gabriele, 2013;Wallace et al, 2016). Results from these immuncytochemistry studies and X-Gal staining experiments in lacZ Eph-ephrin mutants are strikingly consistent with the neuroanatomical patterns described here.…”

“…Experiments pairing modular staining with CRlabeling in the same tissue reveal complementary LCIC zones that are clearly delineated early in development. The described developing neurochemical framework is discussed in light of previous findings from our lab that show similar modular-extramodular patterns revealed by Ephephrin guidance molecule expression (Cramer & Gabriele, 2014;Gabriele et al, 2011;Wallace, Harris, Brubaker, Klotz, & Gabriele, 2016).…”

Alignment of EphA4 and ephrin‐B2 expression patterns with developing modularity in the lateral cortex of the inferior colliculus

“…Much like EphA4, ephrin‐B2 is expressed in the nascent LCIC and exhibits a pattern consistent with that of modular labeling (Cramer & Gabriele, ; Gabriele et al, ; Wallace et al, ). A series of experiments analogous to the presented EphA4 studies was performed for ephrin‐B2 in GAD67‐GFP mice at the same postnatal ages (Figure ).…”

“…These two guidance molecules, while belonging to different subfamilies, are known to exhibit strong binding affinities for each other (Bergemann et al, ; Gale et al, ). Their expression patterns, unlike their characteristic gradients in the neighboring CNIC, are discrete and consist of a discontinuous series of concentrated patches (Cramer & Gabriele, ; Gabriele et al, ; Wallace, Harris, Brubaker, Klotz, & Gabriele, ), reminiscent of the emerging LCIC neurochemical modularity. Despite temporal and spatial similarities in these patterns, it remains unknown how these guidance patterns interface with its developing modular–extramodular framework.…”