A 30 day time course laboratory weathering experiment was conducted using rock samples collected from the West Fork of the Gallatin River watershed (WFW) in southwestern Montana, USA. The goal of these experiments was to quantify the amount of labile nitrogen in rock samples collected from the watershed and determine if chemical weathering is a source of dissolved nitrogen in stream water. Several rock samples investigated produced nitrate in significantly higher concentrations than the silica bead control (p < 0.05), and the data were consistent with elevated NO3− concentrations measured in associated WFW streams. Isotopic analyses of 15N-NO3 in 22 stream water samples from the WFW and four rock:water extracts from the laboratory experiments indicated that the isotopic composition of NO3 was comparable with rocks and stream water samples in the same watershed and differed strongly from waters downstream of development. We suggest that the NO3− measured in WFW streams includes nitrogen derived from mineral dissolution products from soils and rock. The results presented herein further indicate that rock weathering is a source of stream water N in the West Fork watershed and inform water quality assessment, total maximum daily load development, and the relative influences of natural and anthropocentrically derived N sources across this developing mountain watershed.