Our understanding of pain signaling at the spinal level has been greatly advanced during the last decade, both with regard to anatomy, neurochemistry, circuitry and physiology [72–77]. Here we focus on the SST system. Our results provide further evidence that in mouse sst2A is a membrane-bound receptor expressed in a subpopulation of nociceptive DRG neurons and in local dorsal horn neurons. We show that a systematically administrated sst2 agonist, Oct, causes a rapid, transient receptor internalization in DRG neurons as well as counteracts nerve injury-induced pain behaviors in the SNI model, in parallel with attenuation of p-p38 upregulation. sst2A is also internalized in dorsal horn neurons by Oct after ith administration. In some DRG neurons sst2A and the Y1R are co-internalized after Oct stimulation in vivo, hypothetically forming a heterodimer. The sst2A is anterogradely transported and, in fact much more pronounced, also retrogradely, but in this case as a complex with SST, possibly carrying information to the soma. Our findings of sst2A in human DRGs and, abundantly, in spinal cord, suggest that similar mechanisms may operate in rodents and humans, and that targeting sst2A receptors may lead to novel treatment strategies for neuropathic pain.
sst2A in the dorsal horn
Presence of sst2A+ cell bodies and a dense plexus of processes has been reported in the dorsal horn of mouse  and rat [43, 44, 78]. In rat sst2A is present in 13-15% of all neurons in laminae I and II, all are GABA+ and >80% are glycine+, i.e. inhibitory, and they are different from the NK1+
[47, 78] and MOR+
[43, 44] neurons. We here report that many sst2A+ neurons co-express nNOS, a few galanin, SST or NPYY1R, but none PKCgamma. The SST neurons are, on the other hand, glutamatergic, that is excitatory .
Here we show that sst2A is present both in cell bodies in the dorsal horn and in motoneurons in the human spinal cord. In both locations the receptor is mainly membrane-bound. In addition there is a dense plexus of processes in the superficial dorsal horn with a lower density in other layers. The distribution of SST in human DRGs and spinal cord has previously been described [80–83].
Trafficking of sst2A
Trafficking of SST receptors has been studied [84–87]. Here we confirm that i.v. administration of Oct induce internalization of sst2A in DRG neurons . Interestingly, in the latter study sst2A internalization was not observed after treatment with the pan-sst agonist pasireotide, which binds with high affinity to all SSTRs [88, 89].
The sst2A is an endosomally recycled receptor [90, 91]. In the DRG neurons the internalization was almost complete 1 hr after systemic Oct injection, partly back at the plasma membrane after 6 hours, and virtually completely after 24 hrs. Also Oct applied intrathecally induced distinct internalization in the superficial dorsal horn neurons, as well as in DRG neurons. Whether the latter is due to spread to the DRG cell bodies, or represents an Oct action on the central afferents in the dorsal horn, remains to be studied.
Here we show that the Y1R is co-localized with sst2A on the membrane of some DRG neurons. The sst2 agonist Oct induced a parallel internalization of sst2A and Y1R, supporting existence of a heterodimeric complex, a view that however at this point is hypothetical. In previous studies NPY or NPY agonists did not cause internalization of Y1R in rat DRGs (X. Zhang, Z.Q. Xu and T. Hökfelt unpublished observations). Previously, constitutive heterodimerization of sst2A/sst3 has been reported, the heteromers behaving like sst2A dimers, the sst3 being functionally inactivated . Moreover, heterodimerization of sst2A and the μ-opioid receptor did not distinctly change ligand binding or coupling properties .
Axonal transport of sst2A
sst2A is axonally transported in the sciatic nerve, accumulating around the lesion 10 hrs after a crush, the distal pile up, surprisingly, being much more pronounced, and sharply contrasting the strong proximal/modest distal CGRP accumulation. Interestingly, whereas on the proximal side SST+ and sst2A+ fibers were clearly separated from one another, as is expected in view of lack of coexistence in the DRG neuron cell bodies, the two markers appeared to coexist on the distal side. Thus, SST of peripheral origin may bind to sst2A, internalize and travel retrogradely as a SST/sst2A complex. This is supported by presence of sst2A+ nerve fibers in the skin, the distal pile up of exogenously infused SST-RFP and by the preferential, proximal SST accumulation in sst2-KO mice. The marked reduction of distal SST accumulation in the KO mice strongly suggests that the sst2A receptor is required for this pile up in control mice. This is similar to, e.g. nerve growth factor (NGF), and raises the possibility that the SST/sst2A complex represents a retrograde signal, the role of which has still to be defined. The origin of peripheral SST is potentially manifold: ‘endocrine’ SST in the blood, adjacent SST+ sensory branches and/or blood/immune cells .
SST, its receptors and pain
There is now good evidence for a role of SST in pain signaling at the spinal level (see Introduction), involving DRG and local dorsal horn neurons. Both systems produce SST and certain SSTRs. In DRGs, and almost always in spinal cord, SST and sst2A are expressed in separate neuronal populations, excluding autoreceptor mechanisms.
Using unilateral SNI, a neuropathic pain model, the analgesic, sst2A-selective agonist Oct  significantly increased ipsilateral withdrawal threshold in the von Frey test, and decreased the withdrawal response-duration in the Pin-prick (noxious stimulus) and acetone (cold stimulus) tests for several hours. This is a peripheral action on DRG neurons, since Oct does not penetrate the blood–brain-barrier [20, 48]. Moreover, Oct attenuates swelling and mechanical hyperalgesia in a mouse model of immune-mediated arthritis, an effect not seen in sst2A-KO mice , as well as capsaicin-induced nociceptor activity and nociceptive behavior in vitro and in vivo
SST released from peripheral small diameter fibers can inhibit cross-excitation exerted by release of excitatory transmitters, such as glutamate and substance P, from adjacent primary afferent terminals [96, 97], supported by our demonstration of sst2A expression in peripheral nerve terminals in the skin.
Ith SST has been reported to be pro-nociceptive [98–101], like substance P, even if high, probably toxic SST doses are anti-nociceptive [102, 103]. SST causes outwards currents in lamina II sst2A+ neurons [69, 104]. Yasaka et al.  have further clarified the molecular basis for the mainly pro-nociceptive effects exerted by SST in the spinal cord. They showed that SST produces outward currents in inhibitory GABA+/sst2+ interneurons, causing dis-inhibition. Thus, the SNI-induced upregulation of SST in DRG neurons shown here, and a hypothetical increased SST release, could contribute to increased pain signaling in the dorsal horn. However, we also show that ith Oct causes internalization of sst2A, and this may occur after endogenous SST release in the dorsal horn. This internalization should attenuate dis-inhibition and thus offer protection against pain. Finally, it is likely that sst2A is transported into sensory spinal afferents and here represent presynaptic inhibitory receptors, attenuating release of sensory, excitatory transmitters in the dorsal horn, like substance P, CGRP and glutamate, that is a further hypothetical mechanism involved in defense against pain.
It should be mentioned that the sst4 receptor is of interest in relation to pain, a view advanced in particularly by Szolcsányi and colleagues, based on transgenic mice and the use of an sst4-selective agonist, J-2156. Thus, this compound has analgesic effects in several pain models . Moreover, sst4 KO mice exhibit an impaired defense against inflammation and hyperalgesia . However, it is assumed that these sst4 not necessarily are located on neurons in DRGs or spinal cord . Finally, as mentioned above, sst1 may also play a role in nociception based on its high proportion and significant changes in DRGs in response to nerve injury.
NPY, Y1R and pain
The spinal NPY system has been implicated in pain transmission [105, 106]. NPY is abundantly expressed in local dorsal horn neurons but under normal circumstances not in DRG neurons . The Y1R is located both in rodent DRG and dorsal horn neurons [56, 61, 62]. The localization of both Y1R and sst2A on the cell membrane of DRG neurons may confer increased analgesia, since both sst2A and Y1R are anti-nociceptive [20, 21, 24, 42, 56, 108–110]. To what extent co-internalization influences pain signaling remains to be analyzed. This said, it should be emphasized that the Y1R is expressed in a much larger DRG neuron population than sst2A.
Signal transduction pathways
With regard to mechanism of action, we analyzed expression of p38 MAPK [63, 64], a downstream molecule in the sst2 cascade [67, 68]. Its phosphorylated form, p-p38, was strongly upregulated in the ipsilateral DRG after unilateral SNI, paralleling the decrease in pain threshold, in agreement with previous reports [69–71]. Oct, but not saline or Cyn154806, significantly reduced this upregulation, indicating that the sst2/p-p38 pathway may be involved in control of nociceptive thresholds.