Comparative anatomical distribution of neuronal calcium-binding protein (NECAB) 1 and -2 in rodent and human spinal cord

Abstract

Neuronal calcium-binding protein 1 and -2 (NECAB1/2) localize to multiple excitatory neuron populations in the mouse spinal cord. Here, we analyzed rat and human spinal cord, combining in situ hybridization and immunohistochemistry, complementing newly collated data on mouse spinal cord for direct comparisons. Necab1/2 mRNA transcripts showed complementary distribution in rodent's spinal cord. Multiple-labeling fluorescence histochemistry with neuronal phenotypic markers localized NECAB1 to a dense fiber plexus in the dorsal horn, to neurons mainly in superficial layers and to commissural interneurons in both rodent species. NECAB1-positive (+) motor neurons were only found in mice. NECAB1 distribution in the human spinal cord was similar with the addition of NECAB1-like immunoreactivity surrounding myelinated axons. NECAB2 was mainly present in excitatory synaptic boutons in the dorsal horn of all three species, and often in calbindin-D28k(+) neuronal somata. Rodent ependymal cells expressed calbindin-D28k. In humans, they instead were NECAB2(+) and/or calretinin(+). Our results reveal that the association of NECAB2 to excitatory neuronal circuits in the spinal cord is evolutionarily conserved across the mammalian species investigated so far. In contrast, NECAB1 expression is more heterogeneous. Thus, our study suggests that the phenotypic segregation of NECAB1 and -2 to respective excitatory and inhibitory spinal systems can underpin functional modalities in determining the fidelity of synaptic neurotransmission and neuronal responsiveness, and might bear translational relevance to humans.

Recent advances in our understanding of the organization of dorsal horn neuron populations and their contribution to cutaneous mechanical allodynia

“…For example, in mice, expression of the neuronal calcium-binding protein NECAB1 is used to label the cluster DE-5 in Sathyamurthy et al (2018), and is strongly expressed in the Glut5 cluster in Haring et al (2018). However, NECAB1 is expressed almost exclusively by excitatory neurons in the superficial laminae, excluding the PKCγ + population, in mice, but by half of inhibitory neurons expressing the SOM receptor SST2R and half of excitatory PKCγ + neurons in rats (Zhang et al 2014(Zhang et al , 2016. On the other hand, NECAB2 labels similar populations of neurons in the inner part of lamina II in mice, rats and humans (Zhang et al 2014(Zhang et al , 2016, but is expressed by several clusters defined by Haring et al (2018) and Sathyamurthy et al (2018).…”

“…However, NECAB1 is expressed almost exclusively by excitatory neurons in the superficial laminae, excluding the PKCγ + population, in mice, but by half of inhibitory neurons expressing the SOM receptor SST2R and half of excitatory PKCγ + neurons in rats (Zhang et al 2014(Zhang et al , 2016. On the other hand, NECAB2 labels similar populations of neurons in the inner part of lamina II in mice, rats and humans (Zhang et al 2014(Zhang et al , 2016, but is expressed by several clusters defined by Haring et al (2018) and Sathyamurthy et al (2018). There are also important differences in gene expression in distinct spinal segments and DH territories innervated by hairy and glabrous skin, which may be missed in current transcriptomic clustering of DH neurons.…”

Secretagogin-dependent matrix metalloprotease-2 release from neurons regulates neuroblast migration

“…Eluted fractions from secretagogin coimmunoprecipitates were separated on gradient gels (3-12%) and were in-gel digested. Their LC-MS/MS analysis was performed using an an Ultimate 3000 nanoRSLC (Thermo Scientific) coupled to a high-capacity ion trap mass spectrometer (AmaZon ETD; Bruker Daltonics) via a CaptiveSpray ion source (Bruker) (SI Materials and Methods) (75)(76)(77)(78).…”

The basal interstitial nucleus (BIN) of the cerebellum provides diffuse ascending inhibitory input to the floccular granule cell layer

“…Primary antibodies used for immunohistology were from commercial sources with the exception of a rat monoclonal antibody against Lgi2 (see Table 1 for Immunogen, source, catalogue number, RRID number, and final antibody concentration of each primary antibody). The commercial primary antibodies have been well characterized in previous studies (Gao et al, 2016;Jaarsma et al, 1995;Jaarsma, Dino, Cozzari, & Mugnaini, 1996;Singec, Knoth, Ditter, Volk, & Frotscher, 2004;Simat et al, 2007;Zhang et al, 2016). Validation of labeling specificity in this study is based on the expected distribution pattern compared to these studies.…”