Autophagy generates retrogradely transported organelles: a hypothesis.

Nerve cells require trophic signals transmitted from the nerve terminal via the axon in order to survive and develop normally. As the axon may be more than a meter long, specialised mechanisms are needed to transmit these signals. This involves the retrograde axonal transport of signalling endosomes containing nerve growth factor (NGF) and other synaptically derived molecules. These are large, double membrane multivesicular bodies containing a mixture of all vesicle types seen in the nerve terminal. How this signalling endosome is formed and targeted for retrograde axonal transport, however, remains an open question. Here we show that members of the Rab family of proteins that are retrogradely transported indicate that the signalling endosome contains both early and recycling endosomes. In addition, we show that retrogradely transported labelled antibody to dopamine beta-hydroxylase, a marker for synaptic vesicles, co-localizes within the same signalling endosome as NGF. We further show that LC3, a marker for autophagosomes, is retrogradely transported and associates with retrogradely transported NGF. We propose that neurons have exploited the mechanism of autophagy to engulf a sample of the cytoplasmic contents of the nerve terminal to transport back to the cell body. This sample of cytoplasmic contents relays a reliable snapshot of the totality of signalling events occurring in the nerve terminal at that instant in time.

Retrograde axonal transport of dopamine beta hydroxylase antibodies by neurons in the trigeminal ganglion.

In this study we describe a population of neurons in the adult rat trigeminal ganglion (TG) that express dopamine beta-hydroxylase (DBH) and tyrosine hydroxylase (TH), and transport anti-DBH from their terminals. We have used NGF and NT3 labeled with biotin and anti-p75NTR labeled with FITC to examine the transport of neurotrophins and their receptors by these cells. In both the superior cervical ganglion (SCG) and the TG all neurons that transported anti-DBH transported NGF. While 100% of the DBH positive neurons in the TG also transported NT3, approximately 25% of these neurons in the SCG failed to transport NT3. In the SCG virtually all the neurons transported anti-p75NTR with the neurotrophins while in the TG more than 25% of these neurons failed to transport anti-p75NTR with the neurotrophins. These findings suggest that DBH positive neurons in the TG depend upon target-derived NGF and NT3 for their noradrenergic phenotype.

Activation of protein kinase C inhibits retrograde transport of neurotrophins in mice.

Retrograde axonal transport of neurotrophins from nerve terminal to cell body requires a number of key processes, including internalization of the receptor-neurotrophin complex into vesicles and formation of multivesicular bodies and their transport along the axon. Previous studies have shown that each of these processes can be regulated by kinases. In this study, we looked at the role of protein kinase C (PKC) in retrograde transport by injecting labeled neurotrophins together with relevant pharmacological agents into the eye and measuring the accumulation of radioactivity in the trigeminal and superior cervical ganglia. Inhibitors of PKC, Ro-31-8220 and rottlerin, did not affect the retrograde transport of nerve growth factor (NGF); however, phorbol ester activation of classical and novel PKCs blocked retrograde transport. The effect of phorbol esters was partially reversed by rottlerin and Ro-31-8220. Activation of PKC has been shown to be involved in the disorganization of actin filaments. In this study, we show that Ro-31-8220 reverses growth cone collapse by phorbol 12-myristate 13-acetate and suggest that one of the effects of activating PKC on retrograde transport is to disrupt the actin filaments.

Phosphatidylinositol kinase enzymes regulate the retrograde axonal transport of NT-3 and NT-4 in sympathetic and sensory neurons.

Phosphatidylinositol 3-kinase (PI3-kinase) and phosphatidylinositol 4-kinase (PI4-kinase) enzymes are an important family of signaling molecules that have been implicated in the regulation of intracellular vesicle trafficking. It has previously been shown that PI3-kinase and PI4-kinase enzymes regulate neuronal survival and the retrograde axonal transport of nerve growth factor in sympathetic and sensory neurons. We have extended these studies to examine the role these enzymes play in the regulation of the retrograde axonal transport of neurotrophin-3 (NT-3) and neurotrophin-4 (NT-4) in sympathetic and sensory neurons in vivo. Wortmannin (0.1 nmol/eye), a PI3-kinase and PI4-kinase antagonist, reduced the amount of (125)I-NT-3 retrograde transport in sympathetic neurons by approximately 50% and (125)I-NT-4 in sympathetic neurons by approximately 40% and sensory neurons by approximately 20%. The PI3-kinase antagonist LY294002 (100 nmol/eye) reduced the retrograde axonal transport of (125)I-NT-4 in sympathetic and sensory neurons, and (125)I-NT-3 in sympathetic neurons. Phenylarsine oxide (PAO), a PI4-kinase antagonist, significantly inhibited (125)I-NT-4 retrograde axonal transport in sympathetic and sensory neurons. These results show that wortmannin-sensitive PI3-kinases and PI4-kinases may be involved in NT-3 and NT-4 retrograde axonal transport. The retrograde axonal transport of neurotrophic factors in sympathetic and sensory neurons in vivo appears to depend upon the activation of different receptors and second messenger cascades at the nerve terminal.