Development of manganese-enhanced magnetic resonance imaging of the rostral ventrolateral medulla of conscious rats: Importance of normalization and comparison with other regions of interest

Abstract

Spinally projecting neurons in the rostral ventrolateral medulla (RVLM) are believed to contribute to pathophysiological alterations in sympathetic nerve activity and the development of cardiovascular disease. The ability to identify changes in the activity of RVLM neurons in conscious animals and humans, especially longitudinally, would represent a clinically important advancement in our understanding of the contribution of the RVLM to cardiovascular disease. To this end, we describe the initial development of manganese-enhanced magnetic resonance imaging (MEMRI) for the rat RVLM. Manganese (Mn²⁺ ) has been used to estimate in vivo neuronal activity in other brain regions because of both its paramagnetic properties and its entry into and accumulation in active neurons. In this initial study, our three goals were as follows: (1) to validate that Mn²⁺ enhancement occurs in functionally and anatomically localized images of the rat RVLM; (2) to quantify the dose and time course dependence of Mn²⁺ enhancement in the RVLM after one systemic injection in conscious rats (66 or 33 mg/kg, intraperitoneally); and (3) to compare Mn²⁺ enhancement in the RVLM with other regions to determine an appropriate method of normalization of T₁ -weighted images. In our proof-of-concept and proof-of-principle studies, Mn²⁺ was identified by MRI in the rat RVLM after direct microinjection or via retrograde transport following spinal cord injections, respectively. Systemic injections in conscious rats produced significant Mn²⁺ enhancement at 24 h (p < 0.05). Injections of 66 mg/kg produced greater enhancement than 33 mg/kg in the RVLM and paraventricular nucleus of the hypothalamus (p < 0.05 for both), but only when normalized to baseline scans without Mn²⁺ injection. Consistent with findings from our previous functional and anatomical studies demonstrating subregional neuroplasticity, Mn²⁺ enhancement was higher in the rostral regions of the RVLM (p < 0.05). Together with important technical considerations, our studies support the development of MEMRI as a potential method to examine RVLM activity over time in conscious animal subjects.

Keywords: brainstem; conscious; imaging; in vivo; neuronal activity.