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Single-cell microinjection has been successfully used to deliver exogenous proteins, cDNA constructs, peptides, drugs and particles into transfection-challenged cells. With precisely controlled delivery dosage and timing, microinjection has been used in many studies of primary cultured cells, transgenic animal production, in vitro fertilization and RNA inference. This review discusses the advantages and limits of microinjection as a mechanical delivery method and its applications to attached and suspended cells.

We have previously shown that microinjection of galanin into the arcuate nucleus of hypothalamus (ARC) produced antinociceptive effects in rats (Sun et al., 2003a). In this study, the neural pathway of galanin from ARC to midbrain periaqueductal gray (PAG) in nociceptive modulation was investigated. The hindpaw withdrawal latencies (HWLs) with noxious thermal and mechanical stimulation were assessed by the hotplate and the Randall Selitto tests. Intra-ARC administration of 0.1, 0.5, or 1 nmol of galanin induced significant increases in HWLs of rats. The galanin-induced increases in HWLs were inhibited by injection of 10 μg of the opioid receptor antagonist naloxone or 1 nmol of the mu-opioid receptor antagonist beta-funaltrexamine (beta-FNA) into PAG, suggesting that the antinociceptive effects induced by intra-ARC injection of galanin occur via the neural pathway from ARC to PAG. Furthermore, our results demonstrate that the galaninergic fibers directly innervated the beta-endorphinergic neurons in ARC by immunofluorescent methods. Taken together, our results suggest that galanin produces antinociceptive effects in the ARC of rats by activating the beta-endorphinergic pathway from ARC to PAG. © 2007 Wiley-Liss, Inc.

Massage-like stroking induces acute antinociceptive effects that can be reversed by an oxytocin antagonist, indicating activation of oxytocin on endogenous pain controlling systems. We now demonstrate an increase in hindpaw withdrawal latencies (HWLs), in response to thermal and mechanical stimuli, which was present after six treatments of massage-like stroking every other day and which continued to increase through the remaining seven treatments. Repeated massage-like stroking also resulted in increased oxytocin-like immunoreactivity (oxytocin-LI) levels in plasma and periaquaductal grey matter (PAG). Furthermore, increases in HWLs were also present after injections of oxytocin into the PAG (0.1, 0.5 and 1.0 nmol). Intra-PAG oxytocin injection of 1 nmol followed by 1 or 20 nmol of naloxone attenuated the increments in HWL. Also, there was a dose-dependent attenuation of the oxytocin-induced antinociceptive effects following intra-PAG injection of the m-opioid antagonist bfunaltrexamine (b-FNA) and the k-opioid antagonist nor-binaltorphimine (nor-BNI) but not the d-antagonist naltrindole. The longterm antinociceptive effects of massage-like stroking may be attributed, at least partly, to the oxytocinergic system and its interaction with the opioid system, especially the m- and the k-receptors in the PAG.

A major goal of research on drug addiction is to develop the effective treatments to deal with the long-term behavioral disorders especially reinstatement induced by the addictive drugs such as opiates, cocaine, and cannabinoid. The molecular mechanisms underlying these substance-related disorders remain unclear so far. Here we used the model of morphine-induced conditioned place preference (CPP) in rats to mimic the progress of drug-taking, withdrawal and relapse in human. The tissue of ventral tegmental area (VTA), one of the most important brain structures associated with abused drug-related disorders, was taken and two-dimensional electrophoresis (2-DE) was performed to analyze and compare the changes of protein expression patterns during the different stages of morphine-induced CPP. First, we found that there were 80 proteins identified to be changed in the process of morphine-induced CPP. Furthermore, as the mitogen-activated protein kinase kinase 1 (MAPKK1) was increased significantly in the stages of establishment and reinstatement, we confirmed the change of activated extracellular signal-regulated kinase (ERK) by Western blotting in VTA tissue and cultured cell. The results demonstrated that the activated MEK-ERK pathway by chronic morphine treatment in VTA was involved in morphine-induced reinstatement. Moreover, inhibition of MEK-ERK pathway by infusion the MEK inhibitor U0126 in VTA blocked the establishment of morphine-induced CPP. The present study found significant changes in a group of protein expressions in VTA during morphine-induced CPP and further confirmed the role of MEK-ERK cell signaling pathway of VTA in morphine addiction.